Compositions in the form of an injectable aqueous solution comprising amylin, an amylin receptor agonist or an amylin analog, and a co-polyamino acid

ABSTRACT

An injectable aqueous solution, of which the pH is from 6.0 to 8.0, comprising at least: a) amylin, an amylin receptor agonist or an amylin analog; b) a co-polyamino acid bearing carboxylate charges and hydrophobic radicals Hy, said co-polyamino acid consisting of glutamic or aspartic units and said hydrophobic radicals Hy having the following formula I: 
     
       
         
         
             
             
         
       
         
         
           
             wherein the composition does not comprise a basal insulin of which the isoelectric point pI is from 5.8 to 8.5. It also relates to a composition wherein it moreover comprises a prandial insulin.

CROSS REFERENCE TO RELATED APPLICATION:

This is a Continuation of application Ser. No. 15/855,496 filed Dec. 27,2017, which claims priority to FR 1761806 filed Dec. 7, 2017, and FR1663438 filed Dec. 27, 2016. The disclosure of the prior applications ishereby incorporated by reference herein in its entirety.

The invention relates to therapies by injection of amylin, amylinreceptor agonist or amylin analog for treating diabetes.

The invention relates to a composition in the form of an injectableaqueous solution, of which the pH is from 6.0 to 8.0, comprising atleast amylin, an amylin receptor agonist or an amylin analog, and aco-polyamino acid bearing carboxylate charges and hydrophobic radicalsaccording to the invention, and to compositions comprising moreover aninsulin (excluding basal insulins of which the isoelectric point pI isfrom 5.8 to 8.5). The invention also relates to pharmaceuticalformulations comprising the compositions according to the invention.Finally, the invention also relates to a use of the co-polyamino acidsbearing carboxylate charges and hydrophobic radicals according to theinvention for stabilizing compositions of amylin, amylin receptoragonist or amylin analog as well as compositions of amylin, amylinreceptor agonist or amylin analog comprising moreover an insulin.

Type 1 diabetes is an autoimmune disease which leads to the destructionof the beta cells of the pancreas. These cells are known to produceinsulin whose primary function is to regulate the use of glucose in theperipheral tissues (Gerich 1993 Control of glycaemia). Consequently,type 1 diabetes patients suffer from chronic hyperglycemia and have toself-administer exogenous insulin in order to limit this hyperglycemia.Insulin therapy has made it possible to drastically change the lifeexpectancy of these patients. However, the glycemia control ensured byexogenous insulin is not optimal, in particular after the ingestion of ameal. This is connected with the fact that these patients produceglucagon after the ingestion of a meal, which leads to the removal fromstorage of a portion of the glucose stored in the liver, which is notthe case in healthy persons. This glucagon-mediated production ofglucose worsens the problem of glycemia regulation in these patients.

It has been demonstrated that amylin, another hormone produced by thebeta cells of the pancreas and thus also deficient in type 1 diabetespatients, plays a key role in the regulation of post-prandial glycemia.Amylin, also known under the name of “islet amyloid polypeptide” orIAPP, is a 37 aminoacids peptide which is co-stored and co-secreted withinsulin (Schmitz 2004 Amylin Agonists). It is known that this peptideblocks the production of glucagon by the alpha cells of the pancreas.Thus, insulin and amylin have complementary and synergistic functions,since insulin makes it possible to reduce the glucose concentration inthe blood, while amylin makes it possible to reduce the entry ofendogenous glucose into the blood by inhibiting the production(secretion) of endogenous glucagon.

These problems of post-prandial glycemia regulation are quite similarfor type 2 diabetes patients who are treated with insulin to the extentthat their disease has led to a very significant loss of their beta cellmass and consequently of their ability to produce insulin and amylin.

Human amylin has properties which are not compatible with thepharmaceutical requirements in terms of solubility and stability(Goldsbury C S et al., Polymorphic fibrillar assembly of human amylin. JStruct Biol 119: 17-27, 1997). Amylin is known to form amyloid fiberswhich lead to the formation of water-insoluble plaques. Although amylinis the natural hormone, it was necessary to develop an analog in orderto solve these solubility problems.

The physico-chemical properties of amylin thus make its use impossible:amylin is stable for only about fifteen minutes at acidic pH and forless than a minute at neutral pH.

The company Amylin developed an amylin analog, pramlintide, in order toremedy the lack of stability of human amylin. This product, which ismarketed under the name of Symlin®, was approved in 2005 by the FDA forthe treatment of type 1 and type 2 diabetes. It has to be administeredby the subcutaneous route three times daily, within the hour precedingthe meal, in order to improve the control of post-prandial glycemia.This peptide is formulated at acidic pH and has been described asforming fibrils when the pH of the solution is higher than 5.5. Analogvariants are described in the patent U.S. Pat. No. 5,686,411.

This analog is thus not satisfactory with regard to stability when aformulation at neutral pH is considered.

To date, no means exists enabling one to stabilize human amylin in orderto make a pharmaceutical product from it. However, it would beadvantageous for the patients to have access to the human form of thisphysiological hormone. It would also be advantageous to be able toformulate an amylin analog or an amylin receptor agonist at neutral pH.

In addition, it would be of interest to be able to mix the amylin, anamylin analog or an amylin receptor agonist in an aqueous solution witha prandial insulin, since these two products are to be administeredbefore the meal. This would additionally mimic the physiology, sincethese two hormones are co-secreted by the beta cells in response to ameal, in order to improve the control of post-prandial glycemia.

However, taking into account the fact that the solutions of prandialinsulins have a pH close to neutrality for reasons having to do withchemical stability, it is not possible to obtain an aqueous solutionwhich meets the pharmaceutical requirements in terms of solubility andstability.

For this reason, the patent application US2016/001002 of the companyROCHE describes a pump containing two separate reservoirs in order tomake possible the co-administration of these two hormones using a singlemedical device. However, this patent does not solve the problem of themixing of these two hormones in a solution, which would make it possibleto administer them with conventional pumps which are already availableon the market and which comprise only one reservoir.

The patent application WO2013067022 of the company XERIS provides asolution to the problem of stability of the amylin and the problem ofits compatibility with insulin by using an organic solvent instead ofwater. The absence of water seems to solve the stability problems, butthe use of an organic solvent raises safety problems in chronic use fordiabetic patients and also problems of compatibility with the usualmedical devices, in terms of the tubing, the seals and the plasticizersused.

The patent application WO2007104786 of the company NOVO NORDISKdescribes a method which makes it possible to stabilize a solution ofpramlintide, which is an amylin analog, and of insulin by the additionof a phospholipid derived from glycerophosphoglycerol, in particulardimyristoyl glycerophosphoglycerol (DMPG). But this solution requiresthe use of large quantities of DMPG, which can pose a local toleranceproblem.

To the knowledge of the applicant, no satisfactory means exists whichwould make it possible to combine a prandial insulin and human amylin,an amylin receptor agonist or an amylin analog in an aqueous solution soas to enable administration using conventional devices.

The acidic formulation pH and the rapid fibril formation impede theobtention of a pharmaceutical formulation at neutral pH based on amylinand pramlintide, but also the combining of amylin or pramlintide withother active pharmaceutical ingredients, in particular peptides orproteins.

The applicant has noted that, surprisingly, the co-polyamino acidsaccording to the invention stabilize compositions of amylin, amylinreceptor agonist or amylin analog at a pH from 6 to 8. In fact,compositions comprising amylin, an amylin receptor agonist or an amylinanalog in combination with a co-polyamino acid according to theinvention have an increased stability over time, which is of greatinterest for pharmaceutical development.

The applicant also observed that the co-polyamino acids according to theinvention moreover make it possible to obtain a composition comprisingprandial insulin and amylin, amylin receptor agonist or amylin analog,said composition being clear and having an improved stability withregard to fibril formation.

A conventional method for measuring the stabilities of the proteins orpeptides consists in measuring the formation of fibrils with the aid ofThioflavin T, also referred to as ThT. This method makes it possible tomeasure, under temperature and stirring conditions which enable anacceleration of the phenomenon, the latency time before the formation offibrils by measuring the increase in fluorescence. The compositionsaccording to the invention have a latency time before the formation offibrils which is clearly greater than that of amylin, an amylin receptoragonist or an amylin analog at the pH of interest.

The compositions according to the invention have a physical stability,and possibly a chemical stability, which is satisfactory at the desiredpH.

In an embodiment, the invention relates to a composition in the form ofan injectable aqueous solution, of which the pH is from 6.0 to 8.0,comprising at least:

a) amylin, an amylin receptor agonist or an amylin analog;

b) a co-polyamino acid bearing carboxylate charges and hydrophobicradicals Hy, said co-polyamino acid consisting of glutamic or asparticunits and said hydrophobic radicals Hy having the following formula I:

in which

-   -   GpR is a radical of formula II or II′:

-   -   GpA is a radical of formula III or III′:

-   -   GpC is a radical of formula IV:

-   -   the * indicate the sites of attachment of the different groups;    -   a is a whole number equal to 0 or to 1;    -   b is a whole number equal to 0 or to 1;    -   p is a whole number equal to 1 or 2 and        -   if p is equal to 1 then a is equal to 0 or to 1 and GpA is a            radical of formula III′, and        -   if p is equal to 2 then a is equal to 1, and GpA is a            radical of formula III;    -   c is a whole number equal to 0 or to 1, and if c is equal to 0        then d is equal to 1 or to 2;    -   d is a whole number equal to 0, to 1 or to 2;    -   r is a whole number equal to 0 or to 1, and        -   if r is equal to 0 then the hydrophobic radical of formula I            is attached to the co-polyamino acid via a covalent bond            between a carbonyl of the hydrophobic radical and a nitrogen            atom in N-terminal position of the co-polyamino acid, thus            forming an amide function originating from the reaction of            an amine function in N-terminal position of the precursor of            the co-polyamino acid and an acid function borne by the            precursor of the hydrophobic radical, and        -   if r is equal to 1 then the hydrophobic radical of formula I            is attached to the co-polyamino acid:            -   via a covalent bond between a nitrogen atom of the                hydrophobic radial and a carbonyl of the copolyamino                acid, thus forming an amide function originating from                the reaction of an amine function of the precursor of                the hydrophobic radical and an acid function borne by                the precursor of the co-polyamino acid, or            -   via a covalent bond between a carbonyl of the                hydrophobic radical and a nitrogen atom in N-terminal                position of the co-polyamino acid, thus forming an amide                function originating from the reaction of an acid                function of the precursor of the hydrophobic radical and                an amine function in N-terminal position borne by the                precursor of the co-polyamino acid;    -   R is a radical selected from the group consisting of:        -   a linear or branched divalent alkyl radical comprising 2 to            12 carbon atoms if GpR is a radical of formula II or 1 to 11            carbon atoms if GpR is a radical of formula II′ or II″;        -   a linear or branched divalent alkyl radical comprising 2 to            11 carbon atoms if GpR is a radical of formula II or 1 to 11            carbon atoms if GpR is a radical of formula II′ or II″, 1 to            11 carbon atoms, said alkyl radical bearing one or more            —CONH2 functions, and        -   an unsubstituted ether or polyether radical comprising 4 to            14 carbon atoms and 1 to 5 oxygen atoms;    -   A is a linear or branched alkyl radical comprising 1 to 6 carbon        atoms;    -   B is a linear or branched alkyl radical, optionally comprising        an aromatic ring, comprising 1 to 9 carbon atoms;    -   Cx is a linear or branched monovalent alkyl radical, in which x        indicates the number of carbon atoms and:        -   if p is equal to 1, x is from 11 to 25 (11≤x≤25):        -   if p is equal to 2, x is from 9 to 15(9≤x≤15),    -   the ratio i between the number of hydrophobic radicals and the        number of glutamic or aspartic units being from 0 to 0.5        (0<i≤0.5);    -   when several hydrophobic radicals are borne by a co-polyamino        acid, then they are identical or different,    -   the degree of polymerization DP of glutamic or aspartic units is        from 5 to 250;    -   the free acid functions being in the form of a salt of an alkali        cation selected from the group consisting of Na+ and K+;        characterized in that the composition does not comprise a basal        insulin of which the isoelectric point pI is from 5.8 to 8.5.

In an embodiment, Hy comprises more than 30 carbon atoms.

In an embodiment, the composition is characterized in that the pH isfrom 6.6 to 7.8.

In an embodiment, the composition is characterized in that the pH isfrom 7.0 to 7.8.

In an embodiment, the composition is characterized in that the pH isfrom 6.8 to 7.4.

In the formulas, the * indicate the sites of attachment of the differentelements represented.

In the formulas, the * indicate the sites of attachment of thehydrophobic radicals to the co-polyamino acid. The radicals Hy areattached to the co-polyamino acid via amide functions.

In formulas VII and VIIa, the * indicate the sites of attachment of GpR:

-   to the co-polyamino acid and-   to GpA if a=1 or to GPC if a=0.

In formulas III and III′, the * indicate, from left to rightrespectively, the sites of attachment of GpA:

-   -   to GpR if r=1 or to the co-polyamino acid if r=0 and    -   to GpC.

In formula IV, the * indicates the site of attachment of GpC:

-   -   to GpA if a=1, GpR if r=1 and a=0 or    -   to the co-polyamino acid if r=0 and a=0.

All the attachments between the different groups GpR, GpA, GpL, GpG andGpC are amide functions.

The radicals Hy, GpR, GpA, GpL, GpG and GpC, and D are eachindependently identical or different from one monomeric unit to another.

The compositions in the form of an injectable aqueous solution accordingto the invention are clear solutions.

“Clear solution” is understood to mean compositions which satisfy thecriteria described in the American and European pharmacopoeiasconcerning injectable solutions. In the American pharmacopoeia, thesolutions are defined in part <1151> referring to injection (<1>)(referring to <788> according to USP 35 and specified in <788> accordingto USP 35 and in <787>, <788> and <790> USP 38 (starting from Aug. 1,2014), according to USP 38). In the European pharmacopoeia, theinjectable solutions have to meet the criteria given in sections 2.9.19and 2.9.20.

In an embodiment, the composition is characterized in that saidhydrophobic radicals are selected from the hydrophobic radicals offormula I in which, if p is equal to 1 (p=1) and if x is less than orequal to 14 (x≤14), then r=0 or r=1.

In an embodiment, the composition is characterized in that saidhydrophobic radicals are selected from the hydrophobic radicals offormula I in which, if p is equal to 1 (p=1) and if x is from 15 to 16(15≤x≤16), then r=1.

In an embodiment, the composition is characterized in that saidhydrophobic radicals are selected from the hydrophobic radicals offormula I in which, if p is equal to 1 (p=1) and if x is greater than 17(17≤x), then r=1 and R is an ether or polyether radical.

In an embodiment, the composition is characterized in that saidhydrophobic radicals are selected from the hydrophobic radicals offormula I in which, if p is equal to 1 (p=1), then x is from 17 to 25(17≤x≤25).

In an embodiment, the composition is characterized in that thehydrophobic radicals are selected from the hydrophobic radicals offormula I in which p=1, represented by the following formula V:

GpR, GpA, GpC, r and a have the definitions given above.

In an embodiment, the composition is characterized in that thehydrophobic radical is a radical of formula V in which: r is equal to 1(r=1) and a is equal to 0 (a=0).

In an embodiment, the composition is characterized in that thehydrophobic radical is a radical of formula V in which r is equal to 1(r=1) and a is equal to 1 (a=1).

In an embodiment, the composition is characterized in that thehydrophobic radical is a radical of formula V in which GpR is a radicalof formula II.

In an embodiment, the composition is characterized in that thehydrophobic radical is a radical of formula V in which GpR is a radicalof formula II in which R is a divalent linear alkyl radical comprising 2to 12 carbon atoms.

In an embodiment, the composition is characterized in that thehydrophobic radical is a radical of formula V in which GpR is a radicalof formula II in which R is a divalent alkyl radical comprising 2 to 6carbon atoms.

In an embodiment, the composition is characterized in that thehydrophobic radical is a radical of formula V in which GpR is a radicalof formula II in which R is a divalent linear alkyl radical comprising 2to 6 carbon atoms.

In an embodiment, the composition is characterized in that thehydrophobic radical is a radical of formula V in which GpR is a radicalof formula II in which R is a divalent alkyl radical comprising 2 to 4carbon atoms.

In an embodiment, the composition is characterized in that thehydrophobic radical is a radical of formula V in which GpR is a radicalof formula II in which R is a divalent linear alkyl radical comprising 2to 4 carbon atoms.

In an embodiment, the composition is characterized in that thehydrophobic radical is a radical of formula V in which GpR is a radicalof formula II in which R is a divalent alkyl radical comprising 2 carbonatoms.

In an embodiment, the composition is characterized in that thehydrophobic radical is a radical of formula V in which GpR is a radicalof formula II′.

In an embodiment, the composition is characterized in that thehydrophobic radical is a radical of formula V in which GpR is a radicalof formula II′ in which R is a divalent linear alkyl radical comprising1 to 11 carbon atoms.

In an embodiment, the composition is characterized in that thehydrophobic radical is a radical of formula V in which GpR is a radicalof formula II′ in which R is a divalent alkyl radical comprising 1 to 6carbon atoms.

In an embodiment, the composition is characterized in that thehydrophobic radical is a radical of formula V in which GpR is a radicalof formula II or II′, in which R is a divalent alkyl radical, comprising2 to 5 carbon atoms and bearing one or more amide functions (—CONH₂).

In an embodiment, the composition is characterized in that thehydrophobic radical is a radical of formula V in which GpR is a radicalof formula II′ or II, in which R is a divalent linear alkyl radical,comprising 2 to 5 carbon atoms and bearing one or more amide functions(—CONH₂).

In an embodiment, the composition is characterized in that thehydrophobic radical is a radical of formula V in which GpR is a radicalof formula II or II′ in which R is a radical selected from the groupconsisting of the radicals represented by the formulas below:

In an embodiment, the composition is characterized in that the radical Ris attached to the co-polyamino acid via an amide function borne by thecarbon in delta or epsilon position (or in position 4 or 5) with respectto the amide function (—CONH₂).

In an embodiment, the composition is characterized in that thehydrophobic radical is a radical of formula V in which GpR is a radicalof formula II or II′, in which R is an unsubstituted linear ether orpolyether radical comprising 4 to 14 carbon atoms and 1 to 5 oxygenatoms.

In an embodiment, the composition is characterized in that thehydrophobic radical is a radical of formula V in which GpR is a radicalof formula II or II′, in which R is an ether radical.

In an embodiment, the composition is characterized in that thehydrophobic radical is a radical of formula V in which GpR is a radicalof formula II or II′, in which R is an ether radical comprising 4 to 6carbon atoms.

In an embodiment, the composition is characterized in that thehydrophobic radical is a radical of formula V in which GpR is a radicalof formula II or II′ in which R is an ether radical represented by theformula

In an embodiment, the composition is characterized in that thehydrophobic radical is a radical of formula V in which GpR is a radicalof formula II or II′, in which R is a polyether radical.

In an embodiment, the composition is characterized in that thehydrophobic radical is a radical of formula V in which GpR is a radicalof formula II or II′, in which R is a linear polyether radicalcomprising 6 to 10 carbon atoms and 2 to 3 oxygen atoms.

In an embodiment, the composition is characterized in that thehydrophobic radical of formula V in which GpR is a radical of formula IIor II′, in which R is a polyether radical selected from the groupconsisting of the radicals represented by the formulas below:

In an embodiment, the composition is characterized in that thehydrophobic radical of formula V in which GpR is a radical of formula IIin which R is a polyether radical selected from the group consisting ofthe radicals represented by the formulas below:

In an embodiment, the composition is characterized in that thehydrophobic radical is a radical of formula V in which a is equal to 0(a=0) and r is equal to 0 (r=0).

In an embodiment, the composition is characterized in that thehydrophobic radical is a radical of formula V in which a is equal to 1(a=1) and the radical GpA is a radical of formula III′ in which A isselected from the group consisting of the radicals represented by theformulas below:

In an embodiment, the composition is characterized in that thehydrophobic radical is a radical of formula V in which the radical GpCof formula IV is selected from the group consisting of the radicals offormula IVa, IVb or IVc represented hereafter:

In an embodiment, the composition is characterized in that thehydrophobic radical is a radical of formula V in which the radical GpCis of formula IVa.

In an embodiment, the composition is characterized in that thehydrophobic radical is a radical of formula V in which the radical GpCof formula IV is selected from the group consisting of the radicals offormula IVa, IVb or IVc in which b is equal to 0, corresponding toformulas IVd, IVe and IVf, respectively, represented hereafter:

In an embodiment, the composition is characterized in that thehydrophobic radical is a radical of formula V in which the radical GpCcorresponds to formula IV or IVa in which b=0 and to formula IVd.

In an embodiment, the composition is characterized in that thehydrophobic radical is a radical of formula V in which the radical GpCof formula IV in which b=1 is selected from the group consisting of theradicals in which B is an amino acid residue selected from the groupconsisting of the radicals represented by the formulas below:

In an embodiment, the composition is characterized in that thehydrophobic radical is a radical of formula V in which the radical GpCof formula IV or IVa in which b=1 is selected from the group consistingof the radicals in which B is an amino acid residue selected from thegroup consisting of the radicals represented by the formulas below:

In an embodiment, the composition is characterized in that thehydrophobic radical is a radical of formula V in which the radical GpCof formula IV is selected from the group consisting of the radicals inwhich Cx is selected from the group consisting of the linear alkylradicals.

In an embodiment, the composition is characterized in that thehydrophobic radical is a radical of formula V in which the radical GpCof formula IV is selected from the group consisting of the radicals inwhich Cx is selected from the group consisting of the branched alkylradicals.

In an embodiment, the composition is characterized in that thehydrophobic radical is a radical of formula V in which the radical GpCof formula IV is selected from the group consisting of the radicals inwhich Cx is selected from the group consisting of the alkyl radicalscomprising from 11 to 14 carbon atoms.

In an embodiment, the composition is characterized in that thehydrophobic radical is a radical of formula V in which the radical GpCof formula IV is selected from the group consisting of the radicals inwhich Cx is selected from the group consisting of the radicalsrepresented by the formulas below:

In an embodiment, the composition is characterized in that thehydrophobic radical is a radical of formula V in which the radical GpCof formula IV is selected from the group consisting of the radicals inwhich Cx is selected from the group consisting of the alkyl radicalscomprising from 15 to 16 carbon atoms.

In an embodiment, the composition is characterized in that thehydrophobic radical is a radical of formula V in which the radical GpCof formula IV is selected from the group consisting of the radicals inwhich Cx is selected from the group consisting of the radicalsrepresented by the formulas below:

In an embodiment, the composition is characterized in that thehydrophobic radical is a radical of formula V in which the radical GpCof formula IV is selected from the group consisting of the radicals inwhich Cx is selected from the group consisting of the radicalsrepresented by the formulas below:

In an embodiment, the composition is characterized in that thehydrophobic radical is a radical of formula V in which the radical GpCof formula IV is selected from the group consisting of the radicals inwhich Cx is selected from the group consisting of the alkyl radicalscomprising from 17 to 25 carbon atoms.

In an embodiment, the composition is characterized in that thehydrophobic radical is a radical of formula V in which the radical GpCof formula IV is selected from the group consisting of the radicals inwhich Cx is selected from the group consisting of the alkyl radicalscomprising from 17 to 18 carbon atoms.

In an embodiment, the composition is characterized in that thehydrophobic radical is a radical of formula V in which the radical GpCof formula IV is selected from the group consisting of the radicals inwhich Cx is selected from the group consisting of the alkyl radicalsrepresented by the formulas below:

In an embodiment, the composition is characterized in that thehydrophobic radical is a radical of formula V in which the radical GpCof formula IV is selected from the group consisting of the radicals inwhich Cx is selected from the group consisting of the alkyl radicalscomprising from 18 to 25 carbon atoms.

In an embodiment, the composition is characterized in that thehydrophobic radical is a radical of formula V in which the radical GpCof formula IV is selected from the group consisting of the radicals inwhich Cx is selected from the group consisting of the alkyl radicalsrepresented by the formulas below:

In an embodiment, the composition is characterized in that saidhydrophobic radicals are selected from the hydrophobic radicals offormula I in which a=1 and p=2, represented by the following formula VI:

in which

GpR, GpA, GpC and r have the definitions given above.

In an embodiment, the composition is characterized in that thehydrophobic radical is a radical of formula VI in which GpR is a radicalof formula II.

In an embodiment, the composition is characterized in that thehydrophobic radical is a radical of formula VI in which GpR is a radicalof formula II in which R is a divalent linear alkyl radical comprising 2to 12 carbon atoms.

In an embodiment, the composition is characterized in that thehydrophobic radical is a radical of formula VI in which GpR is a radicalof formula II in which R is a divalent alkyl radical comprising 2 to 6carbon atoms.

In an embodiment, the composition is characterized in that thehydrophobic radical is a radical of formula VI in which GpR is a radicalof formula II in which R is a divalent linear alkyl radical comprising 2to 6 carbon atoms.

In an embodiment, the composition is characterized in that thehydrophobic radical is a radical of formula VI in which GpR is a radicalof formula II in which R is an alkyl radical comprising 2 to 4 carbonatoms.

In an embodiment, the composition is characterized in that thehydrophobic radical is a radical of formula VI in which GpR is a radicalof formula II in which R is a divalent linear alkyl radical comprising 2to 4 carbon atoms.

In an embodiment, the composition is characterized in that thehydrophobic radical is a radical of formula VI in which GpR is a radicalof formula II in which R is a divalent linear alkyl radical comprising 2carbon atoms.

In an embodiment, the composition is characterized in that thehydrophobic radical is a radical of formula VI in which GpR is a radicalof formula IL

In an embodiment, the composition is characterized in that thehydrophobic radical is a radical of formula VI in which GpR is a radicalof formula II′ in which R is a divalent linear alkyl radical comprising1 to 11 carbon atoms.

In an embodiment, the composition is characterized in that thehydrophobic radical is a radical of formula VI in which GpR is a radicalof formula II′ in which R is a divalent alkyl radical comprising 1 to 6carbon atoms.

In an embodiment, the composition is characterized in that thehydrophobic radical is a radical of formula VI in which GpR is a radicalof formula II or II′ in which R is a divalent alkyl radical comprising 2to 5 carbon atoms and bearing one or more amide functions (—CONH₂).

In an embodiment, the composition is characterized in that thehydrophobic radical is a radical of formula VI in which GpR is a radicalof formula II or II′ in which R is a divalent linear alkyl radicalcomprising 2 to 5 carbon atoms and bearing one or more amide functions(—CONH₂).

In an embodiment, the composition is characterized in that thehydrophobic radical is a radical of formula VI in which GpR is a radicalof formula II or II′ in which R is a radical selected from the groupconsisting of the radicals represented by the formulas below:

In an embodiment, the composition is characterized in that thehydrophobic radical is a radical of formula VI in which the aminefunction of the radical GpR involved in the formation of the amidefunction which binds said radical GpR to the co-polyamino acid is borneby a carbon in delta or epsilon position (or in position 4 or 5) withrespect to the amide function (—CONH₂).

In an embodiment, the composition is characterized in that thehydrophobic radical is a radical of formula VI in which GpR is a radicalof formula II or II′, in which R is an unsubstituted linear ether orpolyether radical comprising 4 to 14 carbon atoms and 1 to 5 oxygenatoms.

In an embodiment, the composition is characterized in that thehydrophobic radical is a radical of formula VI in which GpR is a radicalof formula II or II′ in which R is an ether radical.

In an embodiment, the composition is characterized in that the etherradical R is a radical comprising 4 to 6 carbon atoms.

In an embodiment, the composition is characterized in that the etherradical is

In an embodiment, the composition is characterized in that thehydrophobic radical is a radical of formula VI in which GpR is a radicalof formula II or II′ in which R is a polyether radical.

In an embodiment, the composition is characterized in that thehydrophobic radical is a radical of formula VI in which GpR is a radicalof formula II or II′ in which R is a linear polyether radical comprising6 to 10 carbon atoms and 2 to 3 oxygen atoms.

In an embodiment, the composition is characterized in that thehydrophobic radical is a radical of formula VI in which GpR is a radicalof formula II or II′ in which R is a linear polyether radical selectedfrom the group consisting of the radicals represented by the formulasbelow:

In an embodiment, the composition is characterized in that thehydrophobic radical is a radical of formula VI in which the radical GpAof formula III is selected from the group consisting of the radicals offormulas IIIa and IIIb represented hereafter:

In an embodiment, the composition is characterized in that thehydrophobic radical is a radical of formula VI in which the radical GpAof formula III is a radical of formula IIIb represented hereafter:

In an embodiment, the composition is characterized in that thehydrophobic radical is a radical of formula VI in which the radical GpCof formula IV is selected from the group consisting of the radicals offormulas IVa, IVb and IVc represented hereafter:

In an embodiment, the composition is characterized in that thehydrophobic radical is a radical of formula VI in which the radical GpCis of formula IVa.

In an embodiment, the composition is characterized in that thehydrophobic radical is a radical of formula VI in which the radical GpCof formula IV is selected from the group consisting of the radicals offormulas IVa, IVb or IVc in which b is equal to 0 (b=0), correspondingto formulas IVd, IVe, and IVf, respectively, represented hereafter:

In an embodiment, the composition is characterized in that thehydrophobic radical is a radical of formula VI in which the radical GpCcorresponds to formula IV or IVa in which b=0 and to formula IVd.

In an embodiment, the composition is characterized in that thehydrophobic radical is a radical of formula VI in which the radical GpCof formula IV is selected from the group consisting of the radicals inwhich Cx is selected from the group consisting of the linear alkylradicals comprising from 9 to 15 carbon atoms.

In an embodiment, the composition is characterized in that thehydrophobic radical is a radical of formula VI in which the radical GpCof formula IV is selected from the group consisting of the radicals inwhich Cx is selected from the group consisting of the branched alkylradicals comprising from 9 to 15 carbon atoms.

In an embodiment, the composition is characterized in that thehydrophobic radical is a radical of formula VI in which the radical GpCof formula IV is selected from the group consisting of the radicals inwhich Cx is selected from the group consisting of the alkyl radicalscomprising 9 or 10 carbon atoms.

In an embodiment, the composition is characterized in that thehydrophobic radical is a radical of formula VI in which the radical GpCof formula IV is selected from the group consisting of the radicals inwhich Cx is selected from the group consisting of the alkyl radicalscomprising from 11 to 15 carbon atoms.

In an embodiment, the composition is characterized in that thehydrophobic radical is a radical of formula VI in which the radical GpCof formula IV is selected from the group consisting of the radicals inwhich Cx is selected from the group consisting of the alkyl radicalscomprising from 11 to 13 carbon atoms.

In an embodiment, the composition is characterized in that thehydrophobic radical is a radical of formula VI in which the radical GpCof formula IV is selected from the group consisting of the radicals inwhich Cx is selected from the group consisting of the radicalsrepresented by the formulas below:

In an embodiment, the composition is characterized in that thehydrophobic radical is a radical of formula VI in which the radical GpCof formula IV is selected from the group consisting of the radicals inwhich Cx is selected from the group consisting of the alkyl radicalscomprising 14 or 15 carbon atoms.

In an embodiment, the composition is characterized in that thehydrophobic radical is a radical of formula VI in which the radical GpCof formula IV is selected from the group consisting of the radicals inwhich Cx is selected from the group consisting of the radicalsrepresented by the formulas below:

In an embodiment, the composition is characterized in that theco-polyamino acid bearing carboxylate charges and hydrophobic radicalsis selected from the co-polyamino acids having the following formulaVII:

in which,

-   -   D represents, independently, either a —CH₂— group (aspartic        unit) or a —CH₂—CH₂— group (glutamic unit),    -   Hy is a hydrophobic radical selected from the hydrophobic        radicals of formula I, V or VI in which r=1 and GpR is a radical        of formula II,    -   R₁ is a hydrophobic radical selected from the hydrophobic        radicals of formula I, V or VI in which r=0 or r=1 and GpR is a        radical of formula II′, or a radical selected from the group        consisting of an H, a linear C2 to C10 acyl group, a branched C3        to C10 acyl group, benzyl, a terminal “amino acid” unit, and a        pyroglutamate,    -   R₂ is a hydrophobic radical selected from the hydrophobic        radicals of formula I, V or VI in which r=1 and GpR is a radical        of formula II, or a —NR′R″ radical, R′ and R″, which may be        identical or different, being selected from the group consisting        of H, the linear or branched or cyclic C2 to C10 alkyls, benzyl,        and said alkyls R′ and R″ optionally forming together one or        more saturated, unsaturated and/or aromatic carbon rings and/or        optionally comprising heteroatoms selected from the group        consisting of O, N and S,    -   X represents an H or a cationic entity selected from the group        comprising the metal cations;    -   n+m represents the degree of polymerization DP of the        co-polyamino acid, that is to say the average number of        monomeric units per co-polyamino acid chain, and 5≤n+m≤250.

The co-polyamino acid bearing carboxylate charges and at least onehydrophobic radical of formula I can also be referred to as“co-polyamino acid” in the present description.

“Statistical co-polyamino acid” refers to a co-polyamino acid bearingcarboxylate charges and at least one hydrophobic radical, a co-polyaminoacid of formula VIIa.

In an embodiment, the composition is characterized in that theco-polyamino acid bearing carboxylate charges and hydrophobic radicalsis selected from the co-polyamino acids of formula VII, in which R₁═R′₁and R₂═R′₂, having the following formula VIIa:

in which,

-   -   m, n, X, D and Hy have the definitions given above;    -   R′₁ is a radical selected from the group consisting of an H, a        linear C2 to C10 acyl group, a branched C3 to C10 acyl group,        benzyl, a terminal “amino acid” unit, and a pyroglutamate;    -   R′₂ is a —NR′R″ radical, R′ and R″, which may be identical or        different, being selected from the group consisting of H, the        linear or branched or cyclic C2 to C10 alkyls, benzyl, and said        alkyls R′ and R″ optionally forming together one or more        saturated, unsaturated and/or aromatic carbon rings and/or        optionally comprising heteroatoms selected from the group        consisting of O, N and S.

“Defined co-polyamino acid” refers to a co-polyamino acid bearingcarboxylate charges and at least one hydrophobic radical, a co-polyaminoacid of formula VIIb.

In an embodiment, the composition is characterized in that theco-polyamino acid bearing carboxylate charges and hydrophobic charges isselected from the co-polyamino acids of formula VII in which n=0 havingthe following formula VIIb:

in which m, X, D, R₁ and R₂ have the definitions given above and atleast R₁ or R₂ is a hydrophobic radical of formula I, V or VI.

In an embodiment, the composition is characterized in that theco-polyamino acid bearing carboxylate charges and hydrophobic charges isselected from the co-polyamino acids of formula VII in which n=0 offormula VIIb and R₁ or R₂ is a hydrophobic radical of formula I, V orVI.

In an embodiment, the composition is characterized in that theco-polyamino acid bearing carboxylate charges and hydrophobic radicalsis selected from the co-polyamino acids of formula VIIb in which R₁ is ahydrophobic radical of formula I, V or VI in which r=0 or r=1 and GpR isof formula II′.

In an embodiment, the composition is characterized in that theco-polyamino acid bearing carboxylate charges and hydrophobic radicalsis selected from the co-polyamino acids of formulas VIIb in which R₂ isa hydrophobic radical of formula I, V or VI in which r=1 and GpR is offormula II.

In an embodiment, the composition is characterized in that R₁ is aradical selected from the group consisting of a linear C₂ to C₁₀ acylradical, a branched C₃ to C₁₀ acyl group, a benzyl, a terminal “aminoacid” unit, and a pyroglutamate.

In an embodiment, the composition is characterized in that R₁ is aradical selected from the group consisting of a linear C₂ to C₁₀ acylgroup or a branched C₃ to C₁₀ acyl group.

In an embodiment, the composition is characterized in that theco-polyamino acid bearing carboxylate charges and hydrophobic radicalsis selected from the co-polyamino acids of formula VII, VIIa or VIIb inwhich the group D is a —CH₂— group (aspartic unit).

In an embodiment, the composition is characterized in that theco-polyamino acid bearing carboxylate charges and hydrophobic radicalsis selected from the co-polyamino acids of formula VII, VIIa or VIIb inwhich the group D is a —CH₂—CH₂— group (glutamic unit).

In an embodiment, the composition is characterized in that the ratio ibetween the number of hydrophobic radicals and the number of glutamic oraspartic acids is from 0.007 to 0.3.

In an embodiment, the composition is characterized in that the ratio ibetween the number of hydrophobic radicals and the number of glutamic oraspartic acids is from 0.01 to 0.3.

In an embodiment, the composition is characterized in that the ratio ibetween the number of hydrophobic radicals and the number of glutamic oraspartic acids is from 0.02 to 0.2.

In an embodiment, the composition is characterized in that thehydrophobic radical corresponds to formula VI and the ratio i betweenthe number of hydrophobic radicals and the number of glutamic oraspartic units is from 0.007 to 0.15.

In an embodiment, the composition is characterized in that thehydrophobic radical corresponds to formula VI and the ratio i betweenthe number of hydrophobic radicals and the number of glutamic oraspartic units is from 0.01 to 0.1.

In an embodiment, the composition is characterized in that thehydrophobic radical corresponds to formula VI and the ratio i betweenthe number of hydrophobic radicals and the number of glutamic oraspartic units is from 0.02 to 0.08.

In an embodiment, the composition is characterized in that thehydrophobic radical corresponds to formula VI in which the radical Cxcomprises between 9 and 10 carbon atoms and the ratio i between thenumber of hydrophobic radicals and the number of glutamic or asparticunits is from 0.03 to 0.15.

In an embodiment, the composition is characterized in that thehydrophobic radical corresponds to formula VI in which the radical Cxcomprises between 11 and 12 carbon atoms and the ratio i between thenumber of hydrophobic radicals and the number of glutamic or asparticunits is from 0.015 to 0.1.

In an embodiment, the composition is characterized in that thehydrophobic radical corresponds to formula VI in which the radical Cxcomprises from 11 to 12 carbon atoms and the ratio i between the numberof hydrophobic radicals and the number of glutamic or aspartic units isfrom 0.02 to 0.08.

In an embodiment, the composition is characterized in that thehydrophobic radical corresponds to formula VI in which the radical Cxcomprises from 13 to 15 carbon atoms and the ratio i between the numberof hydrophobic radicals and the number of glutamic or aspartic units isfrom 0.01 to 0.1.

In an embodiment, the composition is characterized in that thehydrophobic radical corresponds to formula VI in which the radical Cxcomprises from 13 to 15 carbon atoms and the ratio i between the numberof hydrophobic radicals and the number of glutamic or aspartic units isfrom 0.01 to 0.06.

In an embodiment, the composition is characterized in that thehydrophobic radical corresponds to formula V and the ratio i between thenumber of hydrophobic radicals and the number of glutamic or asparticunits is from 0.007 to 0.3.

In an embodiment, the composition is characterized in that thehydrophobic radical corresponds to formula V and the ratio i between thenumber of hydrophobic radicals and the number of glutamic or asparticunits is from 0.01 to 0.3.

In an embodiment, the composition is characterized in that thehydrophobic radical corresponds to formula V and the ratio i between thenumber of hydrophobic radicals and the number of glutamic or asparticunits is from 0.015 to 0.2.

In an embodiment, the composition is characterized in that thehydrophobic radical corresponds to formula V in which the radical Cxcomprises from 11 to 14 carbon atoms and the ratio i between the numberof hydrophobic radicals and the number of glutamic or aspartic units isfrom 0.1 to 0.2.

In an embodiment, the composition is characterized in that thehydrophobic radical corresponds to formula V in which the radical Cxcomprises from 15 to 16 carbon atoms and the ratio i between the numberof hydrophobic radicals and the number of glutamic or aspartic units isfrom 0.04 to 0.15.

In an embodiment, the composition is characterized in that thehydrophobic radical corresponds to formula V in which the radical Cxcomprises from 17 to 18 carbon atoms and the ratio i between the numberof hydrophobic radicals and the number of glutamic or aspartic units isfrom 0.02 to 0.06.

In an embodiment, the composition is characterized in that thehydrophobic radical corresponds to formula V in which the radical Cxcomprises from 19 to 25 carbon atoms and the ratio i between the numberof hydrophobic radicals and the number of glutamic or aspartic units isfrom 0.01 to 0.06.

In an embodiment, the composition is characterized in that thehydrophobic radical corresponds to formula V in which the radical Cxcomprises from 19 to 25 carbon atoms and the ratio i between the numberof hydrophobic radicals and the number of glutamic or aspartic units isfrom 0.01 to 0.05.

In an embodiment, the composition according to the invention ischaracterized in that n+m is from 10 to 250.

In an embodiment, the composition according to the invention ischaracterized in that n+m is from 10 to 200.

In an embodiment, the composition according to the invention ischaracterized in that n+m is from 15 to 150.

In an embodiment, the composition according to the invention ischaracterized in that n+m is from 15 to 100.

In an embodiment, the composition according to the invention ischaracterized in that n+m is from 15to 80.

In an embodiment, the composition according to the invention ischaracterized in that n+m is from 15to 65.

In an embodiment, the composition according to the invention ischaracterized in that n+m is from 20 to 60.

In an embodiment, the composition according to the invention ischaracterized in that n+m is from 20 to 50.

In an embodiment, the composition according to the invention ischaracterized in that n+m is from 20to40.

The invention also relates to said co-polyamino acids bearingcarboxylate charges and hydrophobic radicals of formula I and to theprecursors of said hydrophobic radicals.

The co-polyamino acids bearing carboxylate charges and hydrophobicradicals of formula X are soluble in water distilled at a pH from 6to8,at a temperature of 25° C. and at a concentration of less than 60 mg/mL.

In an embodiment, the invention also relates to the precursors of saidhydrophobic radicals of formula X.

“Soluble” is understood to mean suitable for making it possible toprepare a solution which is clear and free of particles at aconcentration of less than 100 mg/mL in distilled water at 25° C.

“Solution” is understood to mean a liquid solution which is free ofvisible particles, using the method according to the Europeanpharmacopoeia 8.0, in point 2.9.20, and the American pharmacopoeia.

“Physically stable composition” is understood to mean compositionswhich, after a certain storage time at a certain temperature satisfy thecriteria of the visual inspection described in the Europeanpharmacopoeia, the American pharmacopoeia and the internationalpharmacopoeia, that is to say compositions which are clear and whichcontain no visible particles, but also colorless.

“Chemically stable composition” is understood to mean compositionswhich, after storage for a certain time and at a certain temperature,have a minimum recovery of the active principles and which comply withthe specifications applicable to the pharmaceutical products.

“Injectable aqueous solution” is understood to mean water-based solutionwhich meet the conditions of the European and American pharmacopoeiasand which are sufficiently liquid to be injected.

“Co-polyamino acid consisting of glutamic or aspartic units” isunderstood to mean non-cyclic linear chains of glutamic acid or asparticacid units bound to one another by peptide bonds, said chains having aC-terminal portion corresponding to the carboxylic acid at one end, andan N-terminal portion corresponding to the amine at the other end of thechain.

“Alkyl radical” is understood to mean a linear or branched carbon chainwhich comprises no heteroatom.

The co-polyamino acid is a statistical or block co-polyamino acid.

The co-polyamino acid is a statistical co-polyamino acid in the chain ofthe glutamic and/or aspartic units.

In the formulas, the * indicate the sites of attachments of thedifferent elements represented.

In an embodiment, the composition according to the invention ischaracterized in that the co-polyamino acid originates from a polyaminoacid obtained by polymerization.

In an embodiment, the composition according to the invention ischaracterized in that the co-polyamino acid originates from a polyaminoacid obtained by polymerization by ring opening of a derivative ofglutamic acid N-carboxyanhydride or of a derivative of aspartic acidN-carboxyanhydride.

In an embodiment, the composition according to the invention ischaracterized in that the co-polyamino acid originates from a polyaminoacid obtained by polymerization of a derivative of glutamic acidN-carboxyanhydride or of a derivative of aspartic acidN-carboxyanhydride as described in the article Deming, T. J., Adv.Polym. Sci. 2006, 202, 1-18.

In an embodiment, the composition according to the invention ischaracterized in that the co-polyamino acid originates from a polyaminoacid obtained by polymerization of a derivative of glutamic acidN-carboxyanhydride.

In an embodiment, the composition according to the invention ischaracterized in that the co-polyamino acid originates from a polyaminoacid obtained by polymerization of a derivative of glutamic acidN-carboxyanhydride selected from the group consisting of methylpoly-glutamate N-carboxyanhydride (GluOMe-NCA), benzyl poly-glutamateN-carboxyanhydride (GluOBzl-NCA) and t-butyl poly-glutamateN-carboxyanhydride (GluOtBu-NCA).

In an embodiment, the derivative of glutamic acid N-carboxyanhydride ismethyl poly-L-glutamate methyl poly-glutamate N-carboxyanhydride(L-GluOMe-NCA).

In an embodiment, the derivative of glutamic acid N-carboxyanhydride isbenzyl poly-L-glutamate N-carboxyanhydride (L-GluOBzl-NCA).

In an embodiment, the composition according to the invention ischaracterized in that the co-polyamino acid originates from a polyaminoacid obtained by polymerization of a derivative of glutamic acidN-carboxyanhydride or of a derivative of aspartic acidN-carboxyanhydride using as initiator an organometallic complex of atransition metal as described in the publication Deming, T. J., Nature1997, 390, 386-389.

In an embodiment, the composition according to the invention ischaracterized in that the co-polyamino acid originates from a polyaminoacid obtained by polymerization of a glutamic acid N-carboxyanhydride orof a derivative of aspartic acid N-carboxyanhydride using as initiatorammonia or a primary amine as described in the patent FR 2,801,226 andthe references cited by this patent.

In an embodiment, the composition according to the invention ischaracterized in that the co-polyamino acid originates from a polyaminoacid obtained by polymerization of a derivative of glutamic acidN-carboxyanhydride or of a derivative of aspartic acidN-carboxyanhydride using as initiator hexamethyldisilazane as describedin the publication Lu H.; et al., J. Am. Chem. Soc. 2007, 129,14114-14115 or a silylated amine as described in the publication Lu H.;et al., J. Am. Chem. Soc. 2008, 130, 12562-12563.

In an embodiment, the composition according to the invention ischaracterized in that the synthesis method of the polyamino acidobtained by polymerization of a derivative of glutamic acidN-carboxyanhydride or of a derivative of aspartic acidN-carboxyanhydride from which the co-polyamino acid originates comprisesa step of hydrolysis of ester functions.

In an embodiment, this step of hydrolysis of ester functions can consistof a hydrolysis in an acidic medium or of a hydrolysis in a basic mediumor it can be carried out by hydrogenation.

In an embodiment, this step of hydrolysis of ester groups is ahydrolysis in an acidic medium.

In an embodiment, this step of hydrolysis of ester groups is carried outby hydrogenation.

In an embodiment, the composition according to the invention ischaracterized in that the co-polyamino acid originates from a polyaminoacid obtained by depolymerization of a polyamino acid of highermolecular weight.

In an embodiment, the composition according to the invention ischaracterized in that the co-polyamino acid originates from a polyaminoacid obtained by enzymatic depolymerization of a polyamino acid ofhigher molecular weight.

In an embodiment, the composition according to the invention ischaracterized in that the co-polyamino acid originates from a polyaminoacid obtained by chemical depolymerization of a polyamino acid of highermolecular weight.

In an embodiment, the composition according to the invention ischaracterized in that the co-polyamino acid originates from a polyaminoacid obtained by enzymatic and chemical depolymerization of a polyaminoacid of higher molecular weight.

In an embodiment, the composition according to the invention ischaracterized in that the co-polyamino acid originates from a polyaminoacid obtained by depolymerization of a polyamino acid of highermolecular weight selected from the group consisting of sodiumpolyglutamate and sodium polyaspartate.

In an embodiment, the composition according to the invention ischaracterized in that the co-polyamino acid originates from a polyaminoacid obtained by depolymerization of a sodium polyglutamate of highermolecular weight.

In an embodiment, the composition according to the invention ischaracterized in that the co-polyamino acid originates from a polyaminoacid obtained by depolymerization of a sodium polyaspartate of highermolecular weight.

In an embodiment, the composition according to the invention ischaracterized in that the co-polyamino acid is obtained by grafting ahydrophobic group onto a poly-L-glutamic acid or poly-L-aspartic acidusing the methods of amide bond formation which are well known to theperson skilled in the art.

In an embodiment, the composition according to the invention ischaracterized in that the co-polyamino acid is obtained by grafting ahydrophobic group onto a poly-L-glutamic acid or poly-L-aspartic acidusing the methods of amide bond formation used for peptide synthesis.

In an embodiment, the composition according to the invention ischaracterized in that the co-polyamino acid is obtained by grafting ahydrophobic group onto a poly-L-glutamic acid or poly-L-aspartic acid asdescribed in the patent FR 2,840,614.

Amylin, or “islet amyloid polypeptide” (IAPP), is a 37-residue peptidehormone. It is co-secreted with insulin from the pancreatic beta cellsin the ratio of approximately 100/1. Amylin plays a role in glycemicregulation by stopping the secretion of endogenous glucagon and byslowing the gastric emptying and by promoting satiety, thus reducing thepost-prandial glycemic excursions of the blood glucose levels.

The IAPP is treated starting with an 89-residue coding sequence. Theamyloid polypeptide pro-islet (proIAPP, proamylin, proislet protein) isproduced in the pancreatic beta cells (beta cells) in the form of a67-amino acid pro-peptide, 7404 dalton and it undergoespost-translational modifications comprising the protease cleavage toproduce the amylin.

In the present application, the amylin as mentioned refers to thecompounds described in the patents U.S. Pat. Nos. 5,124,314 and5,234,906.

“Analog”, when the term is used to refer to a peptide or a protein, isunderstood to mean a peptide or a protein in which one or moreconstitutive amino acid residues of the primary sequence have beenreplaced by other amino acid residues and/or in which one or moreconstitutive amino acid residues have been eliminated and/or in whichone or more constitutive amino acid residues have been added. Thepercentage of homology allowed for the present definition of an analogis 50%. In the case of amylin, an analog can be derived, for example,from the primary amino acid sequence of amylin by replacing one or morenatural or nonnatural or peptidomimetic amino acids.

“Derivative”, when the term is used to refer to a peptide or a protein,is understood to mean a peptide or a protein or an analog chemicallymodified by a substituent which is not present in the reference peptideor protein or analog, that is to say a peptide or a protein which hasbeen modified by the creation of covalent bonds, in order to introducesubstituents of non-amino acid type.

An agonist of the amylin receptor refers to a compound which imitatesone or more characteristics of the activity of amylin.

Amylin derivatives are described in the article Yan et al., PNAS, vol.103, no. 7, p. 2046-2051, 2006.

In an embodiment, the substituent is selected from the group consistingof fatty chains.

Amylin analogs are described in the patents U.S. Pat. Nos. 5,686,411,6,114,304 or also 6,410,511.

In an embodiment, the composition is characterized in that the amylinanalog is pramlintide (Symlin®) marketed by the company ASTRAZENECA AB.

In an embodiment, the molar ratios co-polyamino acid/amylin, amylinreceptor agonist or amylin analog are from 1.5 to 75.

In an embodiment, the molar ratios co-polyamino acid/amylin, amylinreceptor agonist or amylin analog are from 1.8 to 50.

In an embodiment, the molar ratios co-polyamino acid/amylin, amylinreceptor agonist or amylin analog are from 2 to 35.

In an embodiment, the molar ratios co-polyamino acid/amylin, amylinreceptor agonist or amylin analog are from 2.5 to 30.

In an embodiment, the molar ratios co-polyamino acid/amylin, amylinreceptor agonist or amylin analog are from 3 to 30.

In an embodiment, the molar ratios co-polyamino acid/amylin, amylinreceptor agonist or amylin analog are from 3.5 to 30.

In an embodiment, the molar ratios co-polyamino acid/amylin, amylinreceptor agonist or amylin analog are from 4 to 30.

In an embodiment, the molar ratios co-polyamino acid/amylin, amylinreceptor agonist or amylin analog are from 5 to 30.

In an embodiment, the molar ratios co-polyamino acid/amylin, amylinreceptor agonist or amylin analog are from 7 to 30.

In an embodiment, the molar ratios co-polyamino acid/amylin, amylinreceptor agonist or amylin analog are from 9 to 30.

In an embodiment, the molar ratios co-polyamino acid/amylin are from 3to 75.

In an embodiment, the molar ratios co-polyamino acid/amylin are from 7to 50.

In an embodiment, the molar ratios co-polyamino acid/amylin are from 10to 30.

In an embodiment, the molar ratios co-polyamino acid/amylin are from 15to 30.

In an embodiment, the molar ratios co-polyamino acid/amylin are from 1.5to 75.

In an embodiment, the molar ratios co-polyamino acid/amylin are from 2to 50.

In an embodiment, the molar ratios co-polyamino acid/amylin are from 3to 30.

In an embodiment, the molar ratios co-polyamino acid/amylin are from 4to 30.

In an embodiment, the molar ratios co-polyamino acid/amylin are from 5to 30.

In an embodiment, the molar ratios co-polyamino acid/amylin are from 8to 30.

In an embodiment, the molar ratios co-polyamino acid/amylin are from 10to 30.

In an embodiment, the molar ratios hydrophobic radical Hy/amylin, amylinreceptor agonist or amylin analog are from 1.5 to 150.

In an embodiment, the molar ratios hydrophobic radical Hy/amylin, amylinreceptor agonist or amylin analog are from 1.8 to 100.

In an embodiment, the molar ratios hydrophobic radical Hy/amylin, amylinreceptor agonist or amylin analog are from 2 to 70.

In an embodiment, the molar ratios hydrophobic radical Hy/amylin, amylinreceptor agonist or amylin analog are from 2.5 to 60.

In an embodiment, the molar ratios hydrophobic radical Hy/amylin, amylinreceptor agonist or amylin analog are from 3 to 60.

In an embodiment, the molar ratios hydrophobic radical Hy/amylin, amylinreceptor agonist or amylin analog are from 3.5 to 60.

In an embodiment, the molar ratios hydrophobic radical Hy/amylin, amylinreceptor agonist or amylin analog are from 4 to 60.

In an embodiment, the molar ratios hydrophobic radical Hy/amylin, amylinreceptor agonist or amylin analog are from 5 to 60.

In an embodiment, the molar ratios hydrophobic radical Hy/amylin, amylinreceptor agonist or amylin analog are from 7 to 60.

In an embodiment, the molar ratios hydrophobic radical Hy/amylin, amylinreceptor agonist or amylin analog are from 9 to 60.

In an embodiment, the molar ratios hydrophobic radical Hy/amylin arefrom 5 to 60.

In an embodiment, the molar ratios hydrophobic radical Hy/amylin arefrom 10 to 60.

In an embodiment, the molar ratios hydrophobic radical Hy/amylin arefrom 15 to 60.

In an embodiment, the molar ratios hydrophobic radical Hy/amylin arefrom 1.5 to 60.

In an embodiment, the molar ratios hydrophobic radical Hy/amylin arefrom 2 to 60.

In an embodiment, the molar ratios hydrophobic radical Hy/amylin arefrom 3 to 60.

In an embodiment, the molar ratios hydrophobic radical Hy/amylin arefrom 4 to 60.

In an embodiment, the molar ratios hydrophobic radical Hy/amylin arefrom 5 to 60.

In an embodiment, the molar ratios hydrophobic radical Hy/amylin arefrom 8 to 60.

In an embodiment, the molar ratios hydrophobic radical Hy/amylin arefrom 10 to 60.

In an embodiment, the weight ratios co-polyamino acid/amylin, amylinreceptor agonist or amylin analog are from 1.0 to 70.

In an embodiment, the weight ratios co-polyamino acid/amylin, amylinreceptor agonist or amylin analog are from 1.2 to 45.

In an embodiment, the weight ratios co-polyamino acid/amylin, amylinreceptor agonist or amylin analog are from 1.3 to 30.

In an embodiment, the weight ratios co-polyamino acid/amylin, amylinreceptor agonist or amylin analog are from 1.7 to 27.

In an embodiment, the weight ratios co-polyamino acid/amylin, amylinreceptor agonist or amylin analog are from 2.0 to 27.

In an embodiment, the weight ratios co-polyamino acid/amylin, amylinreceptor agonist or amylin analog are from 2.3 to 27.

In an embodiment, the weight ratios co-polyamino acid/amylin, amylinreceptor agonist or amylin analog are from 2.7 to 27.

In an embodiment, the weight ratios co-polyamino acid/amylin, amylinreceptor agonist or amylin analog are from 3.3 to 27.

In an embodiment, the weight ratios co-polyamino acid/amylin, amylinreceptor agonist or amylin analog are from 4.7 to 27.

In an embodiment, the weight ratios co-polyamino acid/amylin, amylinreceptor agonist or amylin analog are from 6.0 to 27.

In an embodiment, the weight ratios co-polyamino acid/amylin are from2.0 to 67.

In an embodiment, the weight ratios co-polyamino acid/amylin are from4.7 to 27.

In an embodiment, the weight ratios co-polyamino acid/amylin are from6.7 to 27.

In an embodiment, the weight ratios co-polyamino acid/amylin are from 10to 27.

In an embodiment, the weight ratios co-polyamino acid/amylin are from1.0 to 67.

In an embodiment, the weight ratios co-polyamino acid/amylin are from1.3 to 45.

In an embodiment, the weight ratios co-polyamino acid/amylin are from2.7 to 27.

In an embodiment, the weight ratios co-polyamino acid/amylin are from3.3 to 27.

In an embodiment, the weight ratios co-polyamino acid/amylin are from5.3 to 27.

In an embodiment, the weight ratios co-polyamino acid/amylin are from6.7 to 27.

In an embodiment, the composition is characterized in that it moreoverincludes insulin.

In an embodiment, the composition is characterized in that the insulinis a prandial insulin. The prandial insulins are soluble at pH 7.

Prandial insulin is understood to mean a so-called rapid or “regular”insulin.

The so-called rapid prandial insulins are insulins which have to meetthe needs caused by the ingestion of proteins and carbohydrates during ameal; they have to act within less than 30 minutes.

In an embodiment, the so-called “regular” prandial insulin is humaninsulin.

In an embodiment, the prandial insulin is a recombinant human insulin asdescribed in the European pharmacopoeia and the American pharmacopoeia.

The human insulin is marketed, for example, under the trade namesHumulin® (ELI LILLY) and Novolin® (NOVO NORDISK).

The so-called rapid (fast acting) prandial insulins are insulins whichare obtained by recombination and whose primary sequence has beenmodified to decrease their action time.

In an embodiment, the so-called rapid (fast acting) prandial insulinsare selected from the group comprising insulin lispro (Humalog®),insulin glulisine (Apidra®) and insulin aspart (NovoLog®).

In an embodiment, the prandial insulin is insulin lispro.

In an embodiment, the prandial insulin is insulin glulisine.

In an embodiment, the prandial insulin is insulin aspart.

The units recommended by the pharmacopoeias for the insulins arepresented in table 51 below with their equivalents in mg:

TABLE 51 Units recommended by the pharmacopoeias for the insulins USPharmacopoeia - Insulin EP Pharmacopoeia 8.0 (2014) USP38 (2015) Aspart1 U = 0.0350 mg of insulin aspart 1 USP = 0.0350 mg of insulin aspartLispro 1 U = 0.0347 mg of insulin lispro 1 USP = 0.0347 mg of insulinlispro Human 1 IU = 0.0347 mg of human insulin 1 USP = 0.0347 mg ofhuman insulin

In the case of the insulin glulisine, 100 U=3.49 mg of insulin glulisine(according to “Annex 1—Summary of product characteristics” pertaining toAdipra®).

Nevertheless, in the remainder of the text, U is routinely used equallyfor the quantities and the concentrations of all the insulins. Thecorresponding respective values in mg are the values given above forvalues expressed in U, IU or USP.

In an embodiment, it relates to a pharmaceutical formulationcharacterized in that the concentration of insulin is from 240 to 3000μM (40 to 500 U/mL).

In an embodiment, it relates to a pharmaceutical formulationcharacterized in that the concentration of insulin is from 600 to 3000μM (100 to 500 U/mL).

In an embodiment, it relates to a pharmaceutical formulationcharacterized in that the concentration of insulin is from 600 to 2400μM (100 to 400 U/mL).

In an embodiment, it relates to a pharmaceutical formulationcharacterized in that the concentration of insulin is from 600 to 1800μM (100 to 300 U/mL).

In an embodiment, it relates to a pharmaceutical formulationcharacterized in that the concentration of insulin is from 600 to 1200μM (100 to 200 U/mL).

In an embodiment, it relates to a pharmaceutical formulationcharacterized in that the concentration of insulin is 600 μM (100 U/mL).

In an embodiment, it relates to a pharmaceutical formulationcharacterized in that the concentration of insulin is 1200 μM (200U/mL).

In an embodiment, it relates to a pharmaceutical formulationcharacterized in that the concentration of insulin is 1800 μM (300U/mL).

In an embodiment, it relates to a pharmaceutical formulationcharacterized in that the concentration of insulin is 2400 μM (400U/mL).

In an embodiment, it relates to a pharmaceutical formulationcharacterized in that the concentration of insulin is 3000 μM (500U/mL).

In an embodiment comprising prandial insulin, the molar ratiosco-polyamino acid/amylin, amylin receptor agonist or amylin analog arefrom 1.5 to 75.

In an embodiment comprising prandial insulin, the molar ratiosco-polyamino acid/amylin, amylin receptor agonist or amylin analog arefrom 1.8 to 50.

In an embodiment comprising prandial insulin, the molar ratiosco-polyamino acid/amylin, amylin receptor agonist or amylin analog arefrom 2 to 35.

In an embodiment comprising prandial insulin, the molar ratiosco-polyamino acid/amylin, amylin receptor agonist or amylin analog arefrom 2.5 to 30.

In an embodiment comprising prandial insulin, the molar ratiosco-polyamino acid/amylin, amylin receptor agonist or amylin analog arefrom 3 to 30.

In an embodiment comprising prandial insulin, the molar ratiosco-polyamino acid/amylin, amylin receptor agonist or amylin analog arefrom 3.5 to 30.

In an embodiment comprising prandial insulin, the molar ratiosco-polyamino acid/amylin, amylin receptor agonist or amylin analog arefrom 4 to 30.

In an embodiment comprising prandial insulin, the molar ratiosco-polyamino acid/amylin, amylin receptor agonist or amylin analog arefrom 5 to 30.

In an embodiment comprising prandial insulin, the molar ratiosco-polyamino acid/amylin, amylin receptor agonist or amylin analog arefrom 7 to 30.

In an embodiment comprising prandial insulin, the molar ratiosco-polyamino acid/amylin, amylin receptor agonist or amylin analog arefrom 9 to 30.

In an embodiment comprising prandial insulin, the molar ratiosco-polyamino acid/amylin are from 5 to 75.

In an embodiment comprising prandial insulin, the molar ratiosco-polyamino acid/amylin are from 10 to 50.

In an embodiment comprising prandial insulin, the molar ratiosco-polyamino acid/amylin are from 15 to 30.

In an embodiment comprising prandial insulin, the molar ratiosco-polyamino acid/pramlintide are from 1.5 to 75.

In an embodiment comprising prandial insulin, the molar ratiosco-polyamino acid/pramlintide are from 2 to 50.

In an embodiment comprising prandial insulin, the molar ratiosco-polyamino acid/pramlintide are from 3 to 30.

In an embodiment comprising prandial insulin, the molar ratiosco-polyamino acid/pramlintide are from 4 to 30.

In an embodiment comprising prandial insulin, the molar ratiosco-polyamino acid/pramlintide are from 5 to 30.

In an embodiment comprising prandial insulin, the molar ratiosco-polyamino acid/pramlintide are from 8 to 30.

In an embodiment comprising prandial insulin, the molar ratiosco-polyamino acid/pramlintide are from 10 to 30.

In an embodiment comprising prandial insulin, the molar ratioshydrophobic radical Hy/amylin, amylin receptor agonist or amylin analogare from 1.5 to 150.

In an embodiment comprising prandial insulin, the molar ratioshydrophobic radical Hy/amylin, amylin receptor agonist or amylin analogare from 1.8 to 100.

In an embodiment comprising prandial insulin, the molar ratioshydrophobic radical Hy/amylin, amylin receptor agonist or amylin analogare from 2 to 70.

In an embodiment comprising prandial insulin, the molar ratioshydrophobic radical Hy/amylin, amylin receptor agonist or amylin analogare from 2.5 to 60.

In an embodiment comprising prandial insulin, the molar ratioshydrophobic radical Hy/amylin, amylin receptor agonist or amylin analogare from 3 to 60.

In an embodiment comprising prandial insulin, the molar ratioshydrophobic radical Hy/amylin, amylin receptor agonist or amylin analogare from 3.5 to 60.

In an embodiment comprising prandial insulin, the molar ratioshydrophobic radical Hy/amylin, amylin receptor agonist or amylin analogare from 4 to 60.

In an embodiment comprising prandial insulin, the molar ratioshydrophobic radical Hy/amylin, amylin receptor agonist or amylin analogare from 5 to 60.

In an embodiment comprising prandial insulin, the molar ratioshydrophobic radical Hy/amylin, amylin receptor agonist or amylin analogare from 7 to 60.

In an embodiment comprising prandial insulin, the molar ratioshydrophobic radical Hy/amylin, amylin receptor agonist or amylin analogare from 9 to 60.

In an embodiment comprising prandial insulin, the molar ratioshydrophobic radical Hy/amylin are from 5 to 60.

In an embodiment comprising prandial insulin, the molar ratioshydrophobic radical Hy/amylin are from 10 to 60.

In an embodiment comprising prandial insulin, the molar ratioshydrophobic radical Hy/amylin are from 15 to 60.

In an embodiment comprising prandial insulin, the molar ratioshydrophobic radical Hy/pramlintide are from 1.5 to 60.

In an embodiment comprising prandial insulin, the molar ratioshydrophobic radical Hy/pramlintide are from 2 to 60.

In an embodiment comprising prandial insulin, the molar ratioshydrophobic radical Hy/pramlintide are from 3 to 60.

In an embodiment comprising prandial insulin, the molar ratioshydrophobic radical Hy/pramlintide are from 4 to 60.

In an embodiment comprising prandial insulin, the molar ratioshydrophobic radical Hy/pramlintide are from 5 to 60.

In an embodiment comprising prandial insulin, the molar ratioshydrophobic radical Hy/pramlintide are from 8 to 60.

In an embodiment comprising prandial insulin, the molar ratioshydrophobic radical Hy/pramlintide are from 10 to 60.

In an embodiment comprising prandial insulin, the weight ratiosco-polyamino acid/amylin, amylin receptor agonist or amylin analog arefrom 1.0 to 70.

In an embodiment comprising prandial insulin, the weight ratiosco-polyamino acid/amylin, amylin receptor agonist or amylin analog arefrom 1.2 to 45.

In an embodiment comprising prandial insulin, the weight ratiosco-polyamino acid/amylin, amylin receptor agonist or amylin analog arefrom 1.3 to 30.

In an embodiment comprising prandial insulin, the weight ratiosco-polyamino acid/amylin, amylin receptor agonist or amylin analog arefrom 1.7 to 27.

In an embodiment comprising prandial insulin, the weight ratiosco-polyamino acid/amylin, amylin receptor agonist or amylin analog arefrom 2.0 to 27.

In an embodiment comprising prandial insulin, the weight ratiosco-polyamino acid/amylin, amylin receptor agonist or amylin analog arefrom 2.3 to 27.

In an embodiment comprising prandial insulin, the weight ratiosco-polyamino acid/amylin, amylin receptor agonist or amylin analog arefrom 2.7 to 27.

In an embodiment comprising prandial insulin, the weight ratiosco-polyamino acid/amylin, amylin receptor agonist or amylin analog arefrom 3.3 to 27.

In an embodiment comprising prandial insulin, the weight ratiosco-polyamino acid/amylin, amylin receptor agonist or amylin analog arefrom 4.7 to 27.

In an embodiment comprising prandial insulin, the weight ratiosco-polyamino acid/amylin, amylin receptor agonist or amylin analog arefrom 6.0 to 27.

In an embodiment comprising prandial insulin, the weight ratiosco-polyamino acid/amylin are from 3.3 to 67.

In an embodiment comprising prandial insulin, the weight ratiosco-polyamino acid/amylin are from 6.6 to 27.

In an embodiment comprising prandial insulin, the weight ratiosco-polyamino acid/amylin are from 10 to 27.

In an embodiment comprising prandial insulin, the weight ratiosco-polyamino acid/pramlintide are from 1.0 to 67.

In an embodiment comprising prandial insulin, the weight ratiosco-polyamino acid/pramlintide are from 1.2 to 45.

In an embodiment comprising prandial insulin, the weight ratiosco-polyamino acid/pramlintide are from 1.3 to 27.

In an embodiment comprising prandial insulin, the weight ratiosco-polyamino acid/pramlintide are from 1.7 to 27.

In an embodiment comprising prandial insulin, the weight ratiosco-polyamino acid/pramlintide are from 2.0 to 27.

In an embodiment comprising prandial insulin, the weight ratiosco-polyamino acid/pramlintide are from 2.3 to 27.

In an embodiment comprising prandial insulin, the weight ratiosco-polyamino acid/pramlintide are from 2.7 to 27.

In an embodiment comprising prandial insulin, the weight ratiosco-polyamino acid/pramlintide are from 3.3 to 27.

In an embodiment comprising prandial insulin, the weight ratiosco-polyamino acid/pramlintide are from 4.7 to 27.

In an embodiment comprising prandial insulin, the weight ratiosco-polyamino acid/pramlintide are from 6.0 to 27.

Moreover, it is particularly advantageous to combine the amylin, anamylin receptor agonist or an amylin analog, in combination or not witha prandial insulin, with GLP-1, GLP-1 analogs, GLP-1 receptor agonistswhich are routinely referred to as GLP-1 RA. In particular, this makesit possible to potentiate the effect of the insulin and is recommendedin certain diabetes treatment types.

In an embodiment, the GLP-1, GLP-1 analogs or GLP-1 RA are referred toas “rapid.”

“Rapid” is understood to mean GLP-1, GLP-1 analogs or GLP-1 RA whoseapparent half-life of elimination after subcutaneous injection in humansis less than 8 h, in particular less than 5 h, preferably less than 4 hor even less than 3 h, such as, for example, exenatide and lixisenatide.

In an embodiment, the GLP-1, GLP-1 analogs or GLP-1 RA are selected fromthe group consisting of exenatide or Byetta® (ASTRAZENECA), lixisenatideor Lyxumia® (SANOFI), their analogs or derivatives and theirpharmaceutically acceptable salts.

In an embodiment, the GLP-1, GLP-1 analog or GLP-1 RA is exenatide orByetta®, its analogs or derivatives and their pharmaceuticallyacceptable salts.

In an embodiment, GLP-1, GLP-1 analog or GLP-1 RA is lixisenatide orLyxumia®, its analogs or derivatives and their pharmaceuticallyacceptable salts.

In an embodiment, the concentration of exenatide, its analogs orderivatives and their pharmaceutically acceptable salts is in a rangefrom 0.01 to 1.0 per 100 U of insulin.

In an embodiment, the concentration of exenatide, its analogs orderivatives and their pharmaceutically acceptable salts is 0.01 to 0.5mg per 100 U of insulin.

In an embodiment, the concentration of exenatide, its analogs orderivatives and their pharmaceutically acceptable salts is 0.02 to 0.4mg per 100 U of insulin.

In an embodiment, the concentration of exenatide, its analogs orderivatives and their pharmaceutically acceptable salts is 0.03 to 0.3mg per 100 U of insulin.

In an embodiment, the concentration of exenatide, its analogs orderivatives and their pharmaceutically acceptable salts is 0.04 to 0.2mg per 100 U of insulin.

In an embodiment, the concentration of exenatide, its analogs orderivatives and their pharmaceutically acceptable salts is 0.04 to 0.15mg per 100 U of insulin.

In an embodiment, the concentration of lixisenatide, its analogs orderivatives and their pharmaceutically acceptable salts is in a rangefrom 0.01 to 1 mg per 100 U of insulin.

In an embodiment, the concentration of lixisenatide, its analogs orderivatives and their pharmaceutically acceptable salts is in a rangefrom 0.01 to 0.5 mg per 100 U of insulin.

In an embodiment, the concentration of lixisenatide, its analogs orderivatives and their pharmaceutically acceptable salts is 0.02 to 0.4mg per 100 U of insulin.

In an embodiment, the concentration of lixisenatide, its analogs orderivatives and their pharmaceutically acceptable salts is 0.03 to 0.3mg per 100 U of insulin.

In an embodiment, the concentration of lixisenatide, its analogs orderivatives and their pharmaceutically acceptable salts is 0.04 to 0.2mg per 100 U of insulin.

In an embodiment, the concentration of lixisenatide, its analogs orderivatives and their pharmaceutically acceptable salts is 0.04 to 0.15mg per 100 U of insulin.

In an embodiment, the compositions according to the invention areproduced by mixing solutions of amylin and commercial solutions ofGLP-1, GLP-1 analog or GLP-1 receptor agonist RA in volume ratios in arange from 10/90 to 90/10 in the presence of a co-polyamino acid.

In an embodiment, the composition moreover comprises zinc salts.

In an embodiment, the concentration of zinc salts is from 0 to 5000 μM.

In an embodiment, the concentration of zinc salts is from 0 to 4000 μM.

In an embodiment, the concentration of zinc salts is from 0 to 3000 μM.

In an embodiment, the concentration of zinc salts is from 0 to 2000 μM.

In an embodiment, the concentration of zinc salts is from 0 to 1000 μM.

In an embodiment, the concentration of zinc salts is from 50 to 600 μM.

In an embodiment, the concentration of zinc salts is from 100 to 500 μM.

In an embodiment, the concentration of zinc salts is from 200 to 500 μM.

In an embodiment, the zinc salt is zinc chloride.

In an embodiment, the compositions according to the invention moreovercomprise zinc salts at a concentration from 0 to 500 μM per 100 U ofinsulin.

In an embodiment, the compositions according to the invention moreovercomprise zinc salts at a concentration from 0 to 400 μM per 100 U ofinsulin.

In an embodiment, the compositions according to the invention moreovercomprise zinc salts at a concentration from 0 to 300 μM per 100 U ofinsulin.

In an embodiment, the compositions according to the invention moreovercomprise zinc salts at a concentration from 0 to 200 μM per 100 U ofinsulin.

In an embodiment, the compositions according to the invention moreovercomprise zinc salts at a concentration from 0 to 100 μM per 100 U ofinsulin.

In an embodiment, the compositions according to the invention moreovercomprise buffers.

In an embodiment, the compositions according to the invention comprisebuffers at concentrations from 0 to 100 mM.

In an embodiment, the compositions according to the invention comprisebuffers at concentrations from 15 to 50 mM.

In an embodiment, the compositions according to the invention comprise abuffer selected from the group consisting of a phosphate buffer, Tris(trishydroxymethylaminomethane) and sodium citrate.

In an embodiment, the buffer is sodium phosphate.

In an embodiment, the buffer is Tris (trishydroxymethylaminomethane).

In an embodiment, the buffer is sodium citrate.

In an embodiment, the compositions according to the invention moreovercomprise preservatives.

In an embodiment, the preservatives are selected from the groupconsisting of m-cresol and phenol, alone or in a mixture.

In an embodiment, the concentration of preservatives is from 10 to 50mM.

In an embodiment, the concentration of preservatives is from 10 to 40mM.

In an embodiment, the compositions according to the invention moreovercomprise a surfactant.

In an embodiment, the surfactant is selected from the group consistingof propylene glycol and polysorbate.

The compositions according to the invention can moreover compriseadditives such as tonicity agents.

In an embodiment, the tonicity agents are selected from the groupconsisting of glycerol, sodium chloride, mannitol and glycine.

The compositions according to the invention can moreover comprise allthe excipients in compliance with the pharmacopoeias and compatible withthe insulins used at the usual concentrations.

The invention also relates to a pharmaceutical formulation according tothe invention, characterized in that it is obtained by drying and/orlyophilization.

In the case of local and systemic releases, the modes of administrationconsidered are by intravenous, subcutaneous, intradermal orintramuscular route.

The transdermal, oral, nasal, vaginal, ocular, buccal, pulmonary routesof administration are also considered.

The invention also relates to an implantable or transportable pumpcomprising a competition according to the invention.

The invention also relates to the use of a composition according to theinvention intended to be placed in an implantable or transportable pump.

In an embodiment, the pump delivers the composition according to theinvention by means of a bolus, basal flow or a combination of bolus andbasal flow.

In an embodiment, the pump delivers the composition according to theinvention by means of a combination of bolus and basal flow.

In an embodiment, the pump delivering the composition according to theinvention is selected from the group of the following group of pumpreferences: AccuCheck® Combo, Accu-Check® Insight, Accu-Check® Spirit,Animas® 2020, Animas® Vibe, CellNovo, Omnipod®, Minimed® 670G, Minimed®640G, Minimed® 630G, Minimed® 530G, Minimed® Paradigm® Revel, MedtronicParadigm® Veo™, Tandem t:slim®, Tandem t:slim X2®, Tandem t:flex®,Mylife YpsoPump®, BetaBionics iLet®, Asante Snap, Valeritas V-Go®,OneTouch®, Cequr PaQ® and Unilife Imperium®.

In an embodiment, the injection system delivering the compositionaccording to the invention is a so-called “close-loop” or semi“close-loop” injection system.

In an embodiment, the injection system delivering the compositionaccording to the invention is a “close-loop” system, system which isequipped with a processor using an algorithm which takes intoconsideration the quantity of insulin present in the organism of thepatient by estimating this quantity by itself.

In an embodiment, the “close loop” injection system comprises a sensorwhich directly or indirectly supplies the blood glucose level of thepatient, an infusion pump which delivers the product, and a processorwhich receives the measurement of the sensor, calculates the dose ofproduct to be delivered by the pump based on this measurement and on aninternal algorithm which predicts the evolution of the blood glucoselevel and sends the command to the pump to deliver the dose calculated.

In an embodiment, the injection system delivering the compositionaccording to the invention is a semi “close-loop” system, a systemequipped with a processor using an algorithm which takes intoconsideration the quantity of insulin present in the organism of thepatient based on external data.

In an embodiment, this external data is supplied by the patient.

In an embodiment, this external data supplied by the patient concernsthe quantity of carbohydrate ingested by the patient, and the beginningand the end of physical activity.

In an embodiment, this external data supplied by the patient relates tothe quantity of carbohydrate ingested by the patient.

In an embodiment, this external data supplied by the patient concernsthe start and the end of physical activity.

In an embodiment, the algorithm can take into consideration otherexternal data which can be given automatically by a sensor.

In an embodiment, the processor present in the system for injecting thecomposition according to the invention can include additional stepsensuring the safety of the dose administered to the patient.

In an embodiment, the system for injecting the composition according tothe invention comprises a sensor which directly or indirectly suppliesthe blood glucose level of the patient.

In an embodiment, the sensor is selected from the group of the sensorsor sensors equipping the injection systems Medtronic Paradigm® Veo™,MiniMed® 640G with SmartGuard®, Roche Dexcom® G4 PLATINUM CGM, RocheDexcom G5™ Mobile CGM, Abbott Diabetes Care FreeStyle Libre Flashglucose monitoring system, Abbott Diabetes Care FreeStyle Navigator IICGM system.

In an embodiment, the system for injecting the composition according tothe invention comprises a two-compartment pump with a compartmentcontaining the composition according to the invention and a compartmentcontaining another drug.

In an embodiment, the system for injecting the composition according tothe invention comprises a two-compartment pump with a compartmentcontaining the composition according to the invention and a compartmentcontaining glucagon.

In an embodiment, the composition according to the invention can be usedin an artificial pancreas system with several hormones.

In an embodiment, the artificial pancreas system with several hormonesusing the composition according to the invention is a “close-loop” orsemi “close-loop” system.

In an embodiment, the artificial pancreas system comprises thecomposition according to the invention and glucagon as other hormone.

In an embodiment, the artificial pancreas system comprises atwo-compartment pump, one compartment containing the compositionaccording to the invention and one compartment containing glucagon.

In an embodiment, the artificial pancreas system comprising atwo-compartment pump is equipped with a processor using an algorithmwhich calculates two doses, a dose of the composition according to theinvention and a dose of glucagon, the doses calculated by the algorithmbeing sent to the pump and being delivered to the patient.

In an embodiment, the artificial pancreas system comprises two pumps, apump which delivers the composition according to the invention and apump which delivers glucagon.

In an embodiment, the artificial pancreas system comprising two pumpscomprises a processor using an algorithm calculating two doses, a doseof the composition according to the invention and a dose of glucagon,the doses calculated being sent to the pump containing the compositionaccording to the invention and to the second pump containing theglucagon, respectively, the doses calculated by the algorithm beingdelivered to the patient.

In an embodiment, the system for injecting the composition according tothe invention is an intelligent pen.

In an embodiment, the system for injecting the composition according tothe invention is an intelligent pen capable of determining the dose tobe injected.

In an embodiment, the intelligent pen which is capable of determiningthe dose to be injected is equipped with a sensor which directly orindirectly supplies the blood glucose level of the patient.

In an embodiment, the intelligent pen which is capable of determiningthe dose to be injected is equipped with a sensor and with a processorwhich calculates the dose of product to be delivered using themeasurement of the sensor and an algorithm which predicts the evolutionof the blood glucose level and sends the command to the pen to registerthe dose calculated.

In an embodiment, the dose calculated by the processor of theintelligent pen is the dose administered by the patient.

In an embodiment, the intelligent pen is equipped with a processor usingan algorithm taking into consideration the quantity of insulin in theorganism of the patient by estimating itself this quantity or byreceiving this data externally.

In an embodiment, the intelligent pen is equipped with a processor usingan algorithm taking into consideration other external data which can begiven automatically by a sensor or by the patient, such as the quantityof carbohydrate ingested by the patient, or the beginning and the end ofphysical activity.

In an embodiment, the system for injecting the composition according tothe invention comprises a pump, two pumps or an intelligent pen equippedwith a processor using an algorithm.

In an embodiment, the algorithm used by the processor is selected fromthe “PID” (Proportional Integral Derivative) algorithms, of which anexample is described in the article (Ruiz, J. et al., Journal ofdiabetes science and technology, 1123-1130, 2012), the “fuzzy logic”algorithms, of which an example is described in the article (Atlas, E.et al, Diabetes care, 1072-1076, 2010), the “MPC” (Model PredictiveControl) algorithms, of which an example is described in the article(Hovorka, R. et al., Physiol. Meas., 905-920, 2004), and the “PD”(Proportional Derivative) algorithms, of which an example is describedin the article (Jacobs, P. et al, IEEE Trans Biomed Eng, 2569-2581,2014).

The invention also relates to single-dose formulations at a pH from 6.0to 8.0 comprising amylin, an amylin receptor agonist or an amylinanalog, and a co-polyamino acid according to the invention.

The invention also relates to single-dose formulations at a pH from 6.0to 8.0 comprising amylin, an amylin receptor agonist or amylin analog, aco-polyamino acid according to the invention, and a GLP-1, a GLP-1analog or a GLP-1 RA as defined above.

The invention also relates to single-dose formulations at a pH from 6.6to 7.8 comprising amylin, an amylin receptor agonist or an amylinanalog, and a co-polyamino acid according to the invention.

The invention also relates to single-dose formulations at a pH from 6.6to 7.8 comprising amylin, an amylin receptor agonist or an amylinanalog, a co-polyamino acid according to the invention, and a prandialinsulin as defined above.

The invention also relates to single-dose formulations at a pH from 6.6to 7.6 comprising amylin, an amylin receptor agonist or an amylinanalog, and a co-polyamino acid according to the invention.

The invention also relates to single-dose formulations at a pH from 6.6to 7.6 comprising amylin, an amylin receptor agonist or an amylinanalog, a co-polyamino acid according to the invention, and a prandialinsulin as defined above.

In an embodiment, the single-dose formulations moreover comprise aco-polyamino acid as defined above.

In an embodiment, the formulations are in the form of an injectablesolution.

The preparation of a composition according to the invention has theadvantage that it can be prepared by simply mixing an aqueous solutionof amylin, an amylin receptor agonist or an amylin analog, and aco-polyamino acid bearing carboxylate charges and at least onehydrophobic radical according to the invention, in an aqueous solutionor in lyophilized form. If necessary, the pH of the preparation isadjusted to a pH from 6 to 8.

The preparation of a composition according to invention has theadvantage that it can be prepared by simply mixing an aqueous solutionof amylin, an amylin receptor agonist or an amylin analog, prandialinsulin, and a co-polyamino acid bearing carboxylate charges and atleast one hydrophobic radical according to the invention, in an aqueoussolution or in lyophilized form. If necessary, the pH of the preparationis adjusted to a pH from 6 to 8.

In an embodiment, the mixture of prandial insulin and co-polyamino acidis concentrated by ultrafiltration.

If necessary, the composition of the mixture is adjusted in terms ofexcipients such as glycerol, m-cresol, zinc chloride and polysorbate(Tween®) by adding concentrated solutions of these excipients within themixture. If necessary, the pH of the preparation is adjusted to a pHfrom 6 to 8.

In an embodiment, the compositions are characterized in that saidcompositions have a stability measured by ThT greater than the stabilityof a reference composition comprising amylin, an amylin receptor agonistor an amylin analog, but comprising no co-polyamino acid bearingcarboxylate charges and hydrophobic radicals Hy.

In an embodiment, the compositions are characterized in that saidcompositions have a stability measured by ThT greater than the stabilityof a reference composition comprising amylin, an amylin receptor agonistor an amylin analog in combination with an insulin, but comprising noco-polyamino acid bearing carboxylate charges and hydrophobic radicalsHy.

In an embodiment, the compositions are characterized in that saidcompositions have a stability measured by ThT greater than the stabilityof a reference composition comprising amylin, an amylin receptor agonistor an amylin analog in combination with a GLP-1, a GLP-1 analog or aGLP-1 receptor agonist, but comprising no co-polyamino acid bearingcarboxylate charges and hydrophobic radicals Hy.

In an embodiment, the compositions are characterized in that saidcompositions have a stability measured by ThT greater than the stabilityof a reference composition comprising amylin, an amylin receptor agonistor an amylin analog in combination with an insulin or a GLP-1, a GLP-1analog or a GLP-1 receptor agonist, but comprising no co-polyamino acidbearing carboxylate charges and hydrophobic radicals Hy.

The invention also relates to a use of a co-polyamino acid bearingcarboxylate charges and hydrophobic radicals Hy in order to stabilize acomposition comprising amylin, an amylin receptor agonist or an amylinanalog.

The invention also relates to a use of a co-polyamino acid bearingcarboxylate charges and hydrophobic radicals Hy for stabilizing acomposition comprising amylin, an amylin receptor agonist or an amylinanalog and a prandial insulin, and optionally a GLP-1, a GLP-1 analog ora GLP-1 receptor agonist.

The invention relates to a method for stabilizing a compositioncomprising amylin, an amylin receptor agonist or an amylin analog, or toa method for stabilizing a composition comprising amylin, an amylinreceptor agonist or an amylin analog and a prandial analog, andoptionally a GLP-1, a GLP-1 analog or a GLP-1 receptor agonist.

DESCRIPTION OF THE FIGURES

FIG. 1:

This figure is a graphic representation of the determination of thelatency time (LT) by monitoring the fluorescence of Thioflavin T, on acurve with the value of the fluorescence (in a.u. arbitrary units) onthe ordinate and with the time in minutes on the abscissa.

FIG. 2:

FIG. 2 is a representation of the results of the pharmacokinetics ofpramlintide obtained with the compositions described in examples CA1/CA2and CA3. The analysis of these profiles indicates that the compositionof example CA3 comprising the co-polyamino acid BB15, 100 IU/mL ofinsulin and 0.6 mg/mL of pramlintide (curve traced with the squarescorresponding to example CA3) makes it possible to obtain a slowerabsorption of pramlintide than the absorption of the composition of theexample with double injection comprising only pramlintide and insulin(curve traced with the triangles corresponding to the double-injectionexample CA1/CA2).

FIG. 3:

FIG. 3 is a representation of the results of the pharmacokinetics ofpramlintide obtained with the compositions described in examples CA1/CA2and CA4. The analysis of these profiles indicates that the compositionof example CA4 comprising the co-polyamino acid AB24, 100 IU/mL ofinsulin and 0.6 μg/mL of pramlintide (curve traced with the squarescorresponding to example CA4) makes it possible to obtain a slowerabsorption of pramlintide than the absorption of the composition of theexample with double injection comprising only pramlintide and insulin(curve traced with the triangles corresponding to the double-injectionexample CA1/CA2).

The following examples illustrate the invention in a nonlimiting manner.

Part A

AA: Synthesis of the Hydrophobic Molecules in which p=1

The hydrophobic radicals are represented in the following table by thecorresponding hydrophobic molecule before grafting onto the co-polyaminoacid.

TABLE 1A list and structures of the hydrophobic molecules synthesizedaccording to the invention. STRUCTURE OF THE HYDROPHOBIC MOLECULE BEFORENo GRAFTING ONTO THE CO-POLYAMINO ACID AA1

AA2

AA3

AA4

AA5

AA6

AA7

AA8

AA9

AA10

AA11

AA12

AA13

AA14

EXAMPLE AA1 Molecule AA1 Molecule A1: Product Obtained by the ReactionBetween Palmitoyl Chloride and L-proline

A solution of palmitoyl chloride (23.0 g, 83.7 mmol) in acetone (167 mL)is added dropwise in 90 min to a solution of L-proline (10.6 g, 92.1mmol) in aqueous sodium hydroxide 1 N (230 mL, 230 mmol). After 14 h ofstirring at ambient temperature, the heterogeneous mixture is cooled to0° C., then filtered through a sinter filter to yield a white solidwhich is washed with water (2×100 mL), then with diisopropyl ether (100mL). The sodium hydroxide is dried at reduced pressure. The solid isthen dissolved at reflux in 200 mL of water, then 8 mL of a hydrochloricacid solution at 37% are added to obtain pH=1. The opalescent reactionmedium is then cooled to 0° C. The precipitate obtained is filteredthrough a sinter filter, then washed with water (5×50 mL) untilfiltrates at physiological pH from 6.0 to 8.0 are obtained, followed bydrying in an oven at 50° C. under a vacuum overnight. The product ispurified by recrystallization in diisopropyl ether. A white solid isobtained.

Yield: 22.7 g (77%).

¹H NMR (CDCl₃, ppm): 0.88 (3H); 1.19-1.45 (24H); 1.58-1.74 (2H);1.88-2.14 (3H); 2.15-2.54 (3H); 3.47 (1H); 3.58 (1H); 4.41 (0.1H); 4.61(0.9H); 6.60-8.60 (1H).

Molecule A2: Product Obtained by Reaction Between Molecule A1 andBoc-ethylenediamine

To a solution of molecule A1 (75.1 g, 212.4 mmol) in 1500 mL ofchloroform, the following are slowly added successively and at ambienttemperature: N,N-diisopropylethylamine (DIPEA) (68.8 g, 532.3 mmol),1-hydroxybenzotriazole (HOBt) (37.1 g, 274.6 mmol), thenN-(3-dimethylaminopropyl)-N′-ethylcarbodiimide (EDC) (53.1 g, 277.0mmol). After 15 min of stirring at ambient temperature, a solution ofBoc-ethylenediamine (BocEDA) (37.6 g, 234.7 mmol) in 35 mL of chloroformis added. After 18 h of stirring at ambient temperature, an HCl solution0.1 N (2.1 L), then a saturated NaCl solution (1 L) are added. Thephases are separated, then the organic phase is washed successively withan HCl solution 0.1 N/saturated NaCl (2.1 L/1 L), a saturated NaClsolution (2 L), a saturated NaHCO₃ solution (2 L), then a saturated NaClsolution (2 L). The organic phase is dried over anhydrous sodiumsulfate, filtered, then concentrated at reduced pressure. The solidobtained is purified by trituration in diisopropyl ether (3×400 mL), toyield a solid after drying under a vacuum at 40° C.

Yield: 90.4 g (86%).

¹H NMR (CDCl₃, ppm): 0.88 (3H); 1.20-1.37 (24H); 1.44 (9H); 1.54-1.70(2H); 1.79-1.92 (1H); 1.92-2.04 (1H); 2.03-2.17 (1H); 2.17-2.44 (3H);3.14-3.36 (4H); 3.43 (1H); 3.56 (1H); 4.29 (0.1 H); 4.51 (0.9 H); 4.82(0.1H); 5.02 (0.9H); 6.84 (0.1H); 7.22 (0.9H).

Molecule AA1

A hydrochloric acid solution 4 N in dioxane (100 mL, 400 mmol) is addeddropwise and at 0° C. to a solution of molecule A2 (20.1 g, 40.5 mmol)in 330 mL of dichloromethane. After 3 h 30 of stirring at ambienttemperature, the solution is concentrated at reduced pressure. Theresidue is purified by flash chromatography (methanol, dichloromethane)to yield a white solid of molecule AA1 in the form of a hydrochloridesalt.

Yield: 16.3 g (93%).

¹H NMR (CDCl₃, ppm): 0.88 (3H); 1.07-1.40 (24H); 1.49-1.63 (2H);1.77-2.18 (4H); 2.18-2.45 (2H); 3.14-3.32 (2H); 3.42-3.63 (2H);3.63-3.84 (2H); 4.37 (0.1H); 4.48 (0.9H); 6.81-8.81 (4H).

LC/MS (ESI): 396.5; (calculated ([M+H]⁺): 396.4).

EXAMPLE AA2 Molecule AA2 Molecule A3: 15-methylhexadecan-1-ol

Magnesium (9.46 g, 389 mmol) in the form of chips is introduced into athree-neck flask under argon. The magnesium is covered with anhydrousTHF (40 mL), and a few drops of 1-bromo-3-methylbutane are added atambient temperature to initiate the reaction. After the observation ofan exotherm and slight turbidity of the medium, the rest of the1-bromo-3-methylbutane (53.87 g, 357 mmol) is added dropwise in 90 min,while the temperature of the medium remains stable between 50 and 60° C.The reaction mixture is then heated at 70° C. for 2 h.

In a three-neck flask under argon, a solution of 12-bromo-1-dodecanol(43 g, 162.1 mmol) in THF (60 mL) is added dropwise to a solution ofCuCl (482 mg, 4.86 mmol) dissolved in NMP (62 mL) at 0° C. To thissolution, the hot solution of organomanganese prepared extemporaneouslywhile maintaining the temperature of the medium below 20° C. is thenadded dropwise. The mixture is then stirred at ambient temperature for16 h. The medium is cooled to 0° C., and the reaction is stopped byaddition of an aqueous HCl solution 1 N until the pH is 1 and themixture is extracted with ethyl acetate. After washing of the organicphase with a saturated NaCl solution and drying over Na₂SO₄, thesolution is filtered and concentrated under a vacuum to yield an oil.After purification by DCVC on silica gel (cyclohexane, ethyl acetate),an oil which crystallizes at ambient temperature is obtained.

Yield: 32.8 g (74%).

¹H NMR (CDCl₃, ppm): 0.87 (6H); 1.14 (2H); 1.20-1.35 (22H); 1.50-1.55(3H); 3.64 (2H).

Molecule A4: 15-methylhexadecanoic acid

Small portions of potassium permanganate (38.2 g, 241.5 mmol) are addedto a solution of molecule A3 (20.65 g, 80.5 mmol) and tetrabutylammoniumbromide (14.02 g, 42.5 mmol) in a mixture of aceticacid/dichloroethane/water (124/400/320 mL) at ambient temperature. Afterstirring at reflux for 5 h and return to ambient temperature, the mediumis acidified to pH 1 by gradual addition of HCl 5 N. Na₂SO₃ (44.6 g,354.3 mmol) is then added gradually until discoloration of the medium.The aqueous phase is extracted with dichloromethane, and the combinedorganic phases are tried over Na₂SO₄, filtered and concentrated under avacuum. After purification by chromatography on silica gel (cyclohexane,ethyl acetate, acetic acid), a white solid is obtained.

Yield: 19.1 g (quantitative)

¹H NMR (CDCl₃, ppm): 0.87 (6H); 1.14 (2H); 1.22-1.38 (20H); 1.51 (1H);1.63 (2H); 2.35 (2H).

Molecule A5: Product Obtained by Reaction Between Molecule A4 andL-proline

Dicyclohexyl carbodiimide (DCC) (8.01 g, 38.8 mmol) andN-hydroxysuccinimide (NHS) (4.47 g, 38.8 mmol) are added successively toa solution of molecule A4 (10 g, 37 mmol) in THF (360 mL) at 0° C. After17 h of stirring at ambient temperature, the mixture is cooled to 0° C.for 20 min, filtered through a sinter filter. L-Proline (4 g, 37.7mmol), triethylamine (34 mL) and water (30 mL) are added to thefiltrate. After 20 h of stirring at ambient temperature, the medium istreated with an HCl solution −1 N until the pH is 1. The aqueous phaseis extracted with dichloromethane (2×125 mL). The combined organicphases are washed with an aqueous HCl solution 1 N (2×100 mL), water(100 mL), then a saturated aqueous NaCl solution (100 mL). After dryingover Na₂SO₄, the organic phase is filtered, concentrated under a vacuum,and the residue is purified by chromatography on silica gel(cyclohexane, ethyl acetate, acetic acid).

Yield: 9.2 g (72%).

¹H NMR (CDCl₃, ppm): 0.86 (6H); 1.14 (2H); 1.22-1.38 (20H); 1.50 (1H);1.67 (2H); 1.95-2.10 (3H); 2.34 (2H); 2.49 (1H); 3.47 (1H); 3.56 (1H);4.61 (1H).

LC/MS (ESI): 368.3; (calculated ([M+H]⁺): 368.6).

Molecule A6: Product Obtained by Reaction Between Molecule A5 andBoc-ethylenediamine

Triethylamine (TEA) (5.23 mL) and2-(1H-benzotriazol-1-yl)-1,1,3,3-tetramethyluronium tetrafluoroborate(TBTU) at ambient temperature are added to a solution of molecule A5(9.22 g, 25.08 mmol) in a THF/DMF mixture (200/50 mL). After 10 min ofstirring, Boc-ethylenediamine (4.42 g, 27.6 mmol) is added. Afterstirring at ambient temperature for 17 h, the mixture is diluted withwater (300 mL) at 0° C. and stirred at cold temperature for 20 min. Theprecipitate formed is filtered through a sinter filter, and the filtrateis extracted with ethyl acetate. The combined organic phases are washedwith a saturated NaHCO₃ solution, dried over Na₂SO₄, filtered,concentrated under a vacuum, and the residue is purified by flashchromatography (ethyl acetate, methanol).

Yield: 6.9 g (54%).

¹H NMR (CDCl₃, ppm): 0.86 (6H); 1.15 (2H); 1.22-1.38 (20H); 1.43 (9H);1.50 (1H); 1.64 (4H); 1.85 (1H); 1.95 (1H); 2.10 (1H); 2.31 (2H);3.20-3.35 (3H); 3.45 (1H); 3.56 (1H); 4.51 (1H); 5.05 (1H); 7.24 (1H).

LC/MS (ESI): 510.6; (calculated ([M+H]⁺): 510.8).

Molecule AA2

An HCl solution 4 N in dioxane (13 mL) is added to a solution ofmolecule A6 (5.3 g, 10.40 mmol) in dichloromethane (50 mL) at 0° C.After 5 h of stirring at 0° C., the mixture is concentrated under avacuum, dissolved in water, and lyophilized to yield a white solid ofmolecule AA2 in the form of a hydrochloride salt.

Yield: 4.6 g (99%).

¹H NMR (D₂O, ppm): 0.91 (6H); 1.22 (2H); 1.22-1.50 (20H); 1.63 (3H);1.98 (1H); 2.10 (2H); 2.26 (1H); 2.39 (1H); 2.43 (1H); 3.22 (2H);3.45-3.60 (3H); 3.78 (1H); 4.42 (1H).

LC/MS (ESI): 410.4; (calculated ([M+H]⁺): 410.7).

EXAMPLE AA3 Molecule AA3 Molecule A7: Product Obtained by the ReactionBetween Molecule A1 and Boc-tri(ethylene glycol)diamine

By a method similar to the one used for preparing molecule A2 applied tomolecule A1 (4.0 g, 11.3 mmol) and to Boc-tri(ethyleneglycol)diamine(3.1 g, 12.4 mmol), a colorless oil is obtained after purification byflash chromatography (methanol, toluene).

Yield: 5.5 g (84%).

¹H NMR (CDCl₃, ppm): 0.88 (3H); 1.09-1.39 (24H); 1.44 (9H); 1.64 (2H);1.79-2.01 (2H); 2.06-2.43 (4H); 3.23-3.68 (14H); 4.33 (0.2H); 4.56(0.8H); 5.25 (1H); 6.49 (0.2H); 7.13-7.50 (0.8H).

Molecule AA3

By a method similar to the one used for the preparation of molecule AA1applied to molecule A7 (5.5 g, 9.4 mmol), a white solid of molecule AA3in the form of a hydrochloride salt is obtained after purification byflash chromatography (methanol, dichloromethane).

Yield: 4.3 g (92%).

¹H NMR (DMSO-d₆, ppm): 0.85 (3H); 1.08-1.40 (24H); 1.40-1.52 (2H);1.71-2.02 (4H); 2.02-2.31 (2H); 2.90-2.98 (2H); 3.15-3.47 (5H);3.50-3.66 (7H); 4.24 (0.6H); 4.32 (0.4H); 7.83 (0.6H); 7.95 (3H); 8.17(0.4H).

LC/MS (ESI): 484.6; (calculated ([M+H]⁺): 484.4).

EXAMPLE AA4 Molecule AA4 Molecule A8: Product Obtained by the ReactionBetween Molecule A1 and Boc-1-amino-4,7,10-trioxa-13-tridecane amine

By a method similar to the one used for the preparation of molecule A2applied to molecule A1 (4.5 g, 12.7 mmol) and toBoc-1-amino-4,7,10-trioxa-13-tridecane amine (4.5 g, 14.0 mmol), ayellow oil is obtained after purification by flash chromatography(methanol, dichloromethane).

Yield: 7.7 g (92%)

¹H NMR (CDCl₃, ppm): 0.88 (3H); 1.22-1.37 (24H); 1.44 (9H); 1.59-1.67(2H); 1.67-2.00 (6H); 2.06-2.45 (4H); 3.18-3.76 (18H); 4.28 (0.2H); 4.52(0.8H); 4.69-5.04 (1H); 6.77 (0.2H); 7.20 (0.8H).

Molecule AA4

By a method similar to the one used for the preparation of molecule AA1applied to molecule A8 (7.7 g, 11.8 mmol), a yellow oil is obtainedafter purification by flash chromatography (methanol, dichloromethane).A co-evaporation with diisopropyl ether makes it possible to obtainmolecule AA4 in the form of a hydrochloride salt in the form of a whitesolid which is dried under a vacuum at 50° C.

Yield: 5.4 g (76%).

¹H NMR (CDCl₃, ppm): 0.88 (3H); 1.08-1.40 (24H); 1.49-1.65 (2H);1.76-2.39 (10H); 3.07-3.28 (3H); 3.34-3.80 (15H); 4.34 (0.05H); 4.64(0.95H); 7.35 (0.05H); 7.66-8.58 (3.95H).

LC/MS (ESI): 556.7; (calculated ([M+H]⁺): 556.5).

EXAMPLE AA5 Molecule AA5 Molecule A9: Product Obtained by ReactionBetween Molecule A1 and the Methyl Ester of N-Boc-L-lysine

By a method similar to the one used for the preparation of molecule A2applied to molecule A1 (4 g, 11.3 mmol) and to the methyl ester ofN-Boc-L-lysine (3.2 g, 12.4 mmol), a colorless oil is obtained afterpurification by flash chromatography (methanol, dichloromethane).

Yield: 4.9 g (73%).

¹H NMR (CDCl₃, ppm): 0.88 (3H); 0.99-1.54 (37H); 1.54-1.75 (3H);1.75-2.04 (3H); 2.04-2.41 (4H); 2.94-3.19 (2H); 3.19-3.81 (5H);4.28-4.64 (2H); 4.94 (1H); 6.45 (0.1H); 7.36 (0.9H).

LC/MS (ESI): 596.7; (calculated ([M+H]⁺): 596.5)

Molecule A10: Product Obtained by Treatment of Molecule A9 with Ammonia

320 mL of an ammonia solution 7 N in methanol are added to a suspensionof molecule A9 (4.9 g, 8.2 mmol) in 10 mL of methanol. After 19 h ofstirring at ambient temperature in a closed atmosphere, 100 mL ofadditional ammonia solution are added. After 24 h of stirring at ambienttemperature in a closed atmosphere, the reaction medium is concentratedat reduced pressure. The residue is purified by trituration indiisopropyl ether at reflux (100 mL), to yield a white solid which isdried under a vacuum at 50° C.

Yield: 4.1 g (85%).

¹H NMR (CDCl₃, ppm): 0.88 (3H); 1.06-1.57 (37H); 1.57-1.79 (3H);1.88-2.41 (7H); 3.09 (2H); 3.49 (1H); 3.62 (1H); 4.34 (1H); 4.51 (1H);4.69-4.81 (1H); 5.43 (0.95H); 5.57 (0.05H); 6.25 (0.05H); 6.52 (0.95H);6.83 (0.05H); 7.11 (0.95H).

Molecule AA5

By a method similar to the one used for the preparation of molecule AA1applied to molecule A10 (388 mg, 0.67 mmol), a white solid of moleculeAA5 in the form of a hydrochloride salt is obtained after purificationby trituration in diisopropyl ether.

Yield: 292 mg (85%):

¹H NMR (DMSO-d6, ppm): 0.85 (3H); 1.06-2.34 (38H); 2.61-2.81 (2H);3.29-3.68 (2H); 4.05-4.17 (1.7H); 4.42 (0.3H); 7.00 (1H); 7.16 (0.7H);7.43 (0.3H); 7.73-8.04 (3.7H); 8.16 (0.3H).

LC/MS (ESI): 481.6; (calculated ([M+H]⁺): 481.4).

EXAMPLE AA6 Molecule AA6 Molecule A11: Product Obtained by the ReactionBetween stearoyl chloride and L-proline

By a method similar to the one used for the preparation of molecule A1applied to L-proline (5.0 g, 43.4 mmol) and to stearoyl chloride (12.0g, 39.6 mmol), a white solid is obtained after purification by flashchromatography (methanol, dichloromethane).

Yield: 5.37 g (36%).

¹H NMR (CDCl₃, ppm): 0.88 (3H); 1.26-1.37 (28H); 1.64-1.70 (2H);1.88-2.10 (3H); 2.36 (2H); 2.54-2.58 (1H); 3.46 (1H); 3.56 (1H); 4.62(1H).

LC/MS (ESI): 382.6; (calculated ([M+H]⁺): 382.3).

Molecule A12: Product Obtained by Reaction Between Molecule A11 andBoc-tri(ethylene glycol)diamine

By a method similar to the one used for the preparation of molecule A6applied to molecule A11 (33.81 g, 88.6 mmol) and to Boc-tri(ethyleneglycol)diamine (26.4 g, 106.3 mmol) in THF using DIPEA instead of TEA, awhite solid is obtained after purification by flash chromatography(ethyl acetate, methanol).

Yield: 43.3 g (80%).

¹H NMR (CDCl₃, ppm): 0.87 (3H); 1.24 (30H); 1.43 (9H); 1.61 (2H); 1.82(1H); 1.96 (1H); 2.25-2.45 (2H); 3.25-3.65 (14H); 4.30 (0.15H); 4.53(0.85H); 5.25 (1H); 6.43 (0.15H); 7.25 (0.85H).

LC/MS (ESI): 612.6; (calculated ([M+H]⁺): 612.9).

Molecule AA6

By a method similar to the one used for the preparation of molecule AA2applied to molecule A12 (43 g, 70.3 mmol), the residue obtained afterconcentration under a vacuum is triturated in acetonitrile. Thesuspension is filtered, and the solid is washed with acetonitrile thenwith acetone. After drying under a vacuum, a white solid of molecule AA6in the form of a hydrochloride salt is obtained.

Yield: 31.2 g (81%).

¹H NMR (DMSO-d₆, ppm): 0.85 (3H); 1.23 (28H); 1.45 (2H); 1.70-2.05 (4H);2.13 (1H); 2.24 (1H); 2.95 (2H); 3.10-3.25 (2H); 3.30-3.65 (10H);4.20-4.45 (1H); 7.85-8.25 (4H).

LC/MS (ESI): 512.4; (calculated ([M+H]⁺): 512.8).

EXAMPLE AA7 Molecule AA7 Molecule A13: Product Obtained by ReactionBetween arachidonic acid and L-proline

By a method similar to the one used for the preparation of molecule A5applied to arachidonic acid (15.51 g, 49.63 mmol) and to L-proline (6 g,52.11 mmol), using DIPEA instead of TEA, a white solid is obtained afterpurification by chromatography column on silica gel (cyclohexane, ethylacetate, acetic acid).

Yield: 12.9 g (63%).

¹H NMR (CDCl₃, ppm): 0.88 (3H); 1.28 (34H); 1.66 (2H); 1.95-2.15 (2H);2.34 (2H); 2.45 (1H); 3.47 (1H); 3.56 (1H); 4.60 (1H).

LC/MS (ESI): 410.4; (calculated ([M+H]⁺): 410.6).

Molecule A14: Product Obtained by the Reaction Between Molecule A13 andBoc-1-amino-4,7,10-trioxa-13-tridecane

By a method similar to the one used for the preparation of molecule A12applied to molecule A13 (10.96 g, 26.75 mmol) and toBoc-1-amino-4,7,10-trioxa-13-tridecane (10.29 g, 32.11 mmol), a solid isobtained after purification by chromatography column on silica gel(cyclohexane, ethyl acetate, methanol).

Yield: 14.2 g (75%).

¹H NMR (CDCl₃, ppm): 0.88 (3H); 1.24 (32H); 1.43 (9H); 1.61 (2H); 1.80(1H); 1.96 (1H); 2.10-2.45 (4H); 3.20-3.75 (18H); 4.30 (0.20H); 4.55(0.80H); 5.03 (1H); 6.75 (0.20H); 7.20 (0.80H).

LC/MS (ESI): 712.8; (calculated ([M+H]⁺): 713.1).

Molecule AA7

After a method similar to the one used for the preparation of moleculeAA2 applied to molecule A14 (14.25 g, 20.01 mmol), the residue obtainedafter concentration under a vacuum of the reaction medium is dissolvedin methanol and evaporated at reduced pressure, the operation beingrepeated 4 times to yield a white solid of molecule AA7 in the form of ahydrochloride salt.

Yield: 12.7 g (98%).

¹H NMR (DMSO-d₆, ppm): 0.85 (3H); 1.23 (32H); 1.45 (2H); 1.64 (2H);1.70-2.05 (6H); 2.10-2.30 (2H); 2.82 (2H); 3.08 (2H); 3.30-3.60 (15H);4.15-4.30 (1H); 7.73-8.13 (4H).

LC/MS (ESI): 612.7; (calculated ([M+H]⁺): 612.9).

EXAMPLE AA8 Molecule AA8 Molecule A15: Product Obtained by the ReactionBetween L-leucine and palmitoyl chloride

By a method similar to the one used for the preparation of molecule A1applied to L-leucine (15.0 g, 114.4 mmol) and to palmitoyl chloride(34.5 g, 125 mmol), a white solid is obtained by trituration indiisopropyl ether.

Yield: 13.0 g (31%).

¹H NMR (CDCl₃, ppm): 0.88 (3H); 0.96 (6H); 1.16-1.35 (24H); 1.55-1.77(5H); 2.23 (2H); 4.55-4.60 (1H); 5.88 (1H).

Molecule A16: Product Obtained by the Reaction Between Molecule A15 andthe Methyl Ester of L-proline

By a method similar to the one used for the preparation of molecule A2applied to molecule A15 (6.00 g, 16.2 mmol) and to the methyl ester ofL-proline (3.23 g, 19.5 mmol), a slightly yellow oil is obtained afterpurification by flash chromatography (methanol, dichloromethane).

Yield: 5.8 g (74%).

¹H NMR (CDCl₃, ppm): 0.83-1.00 (9H); 1.18-1.32 (24H); 1.40-1.73 (5H);1.84-2.33 (6H); 3.47-3.89 (2H); 3.70 (1.14H); 3.71 (1.21H); 3.74(0.53H); 3.76 (0.12H); 4.40-4.56 (1H); 4.63-4.67 (0.04H); 4.84 (0.38);4.90 (0.40); 5.06 (0.18); 5.99 (0.18H); 6.08-6.21 (0.82).

LC/MS (ESI): 481.6; (calculated ([M+H]⁺): 481.4).

Molecule A17: Product Obtained by the Saponification of the Methyl Esterof Molecule A16

Sodium hydroxide 1 N (13.5 mL, 13.5 mmol) is added to a solution ofmolecule A16 (5.8 g, 12.06 mmol) in 30 mL of methanol. After 20 h ofstirring at ambient temperature, the solution is diluted with water,then acidified with 20 mL of hydrochloric acid 1 N at 0° C. Theprecipitate is filtered, then rinsed with water (50 mL), before beingsolubilized in 50 mL of dichloromethane. The organic phase is dried overNa₂SO₄, filtered, then concentrated at reduced pressure to yield acolorless oil.

Yield: 4.5 g (80%).

¹H NMR (CDCl₃, ppm): 0.85-0.99 (9H); 1.14-1.41 (24H); 1.43-1.72 (5H);1.87-2.47 (7H); 3.48-3.55 (0.6H); 3.56-3.62 (0.4H); 3.83-3.90 (0.4H);3.90-3.96 (0.6H); 4.52-4.56 (0.6H); 4.56-4.59 (0.4H); 4.80-4.86 (0.4H);4.86-4.91 (0.6H); 6.05 (0.4H); 6.11 (0.6H).

LC/MS (ESI): 467.6; (calculated ([M+H]⁺): 467.4).

Molecule A18: Product Obtained by Reacting Boc-ethylenediamine andMolecule A17

By a method similar to the one used for the preparation of molecule A2applied to molecule A17 (4.5 g, 9.64 mmol) and to Boc-ethylenediamine(1.70 g, 10.61 mmol), a colorless oil is obtained after purification byflash chromatography (methanol, dichloromethane).

Yield: 2.0 g (34%)

¹H NMR (CDCl₃, ppm): 0.83-0.99 (9H); 1.19-1.32 (24H); 1.44 (9H);1.48-2.37 (14H); 3.09-3.99 (4H); 4.28-5.01 (2H); 5.64-6.04 (1H);6.87-7.06 (1H).

LC/MS (ESI): 609.7; (calculated ([M+H]⁺): 609.5).

Molecule AA8

By a method similar to the one used for the preparation of molecule AA1applied to molecule A18 (2 g, 3.28 mmol), a solid of molecule AA8 in theform of a hydrochloride salt is obtained after purification by flashchromatography (methanol, dichloromethane).

Yield: 1.5 g (90%).

¹H NMR (CDCl₃, ppm): 0.83-1.00 (9H); 1.18-1.32 (24H); 1.37-1.77 (5H);1.93-2.41 (6H); 3.07-3.97 (6H); 4.44-4.77 (2H); 7.66-8.21 (2H).

LC/MS (ESI): 509.6; (calculated ([M+H]⁺): 509.4).

EXAMPLE AA9 Molecule AA9 Molecule A19: Product Obtained by the ReactionBetween lauric acid and L-phenylalanine

By a method similar to the one used for the preparation of molecule A5applied to lauric acid (8.10 g, 40.45 mmol) and to L-phenylalanine (7 g,42.38 mmol), a white solid is obtained.

Yield: 12.7 g (98%).

¹H NMR (DMSO-d₆, ppm): 0.86 (3H); 1.10-1.30 (16H); 1.36 (2H); 2.02 (2H);2.82 (1H); 3.05 (1H); 4.42 (1H); 7.15-7.30 (5H); 8.05 (1H); 12.61 (1H).

LC/MS (ESI): 348.2; (calculated ([M+H]⁺): 348.5).

Molecule A20: Product Obtained by the Reaction Between Molecule A19 andthe Hydrochloride Salt of the Methyl Ester of L-proline

By a method similar to the one used for the preparation of molecule A6applied to molecule A19 (9.98 g, 28.72 mmol) and to the hydrochloridesalt of the methyl ester of L-proline (5.23 g, 31.59 mmol), a colorlessoil is obtained after purification by chromatography column on silicagel (cyclohexane, ethyl acetate).

Yield: 5.75 g (44%).

¹H NMR (CDCl₃, ppm): 0.88 (3H); 1.10-1.30 (16H); 1.50-1.75 (3H);1.80-2.02 (3H); 2.17 (2H); 2.65 (0.5H); 2.95 (1H); 3.05-3.20 (1.5H);3.50-3.65 (1H); 3.75 (3H); 4.29 (0.5H); 4.46 (0.5H); 4.70 (0.1H); 4.95(0.9H); 6.20-6.30 (1H); 7.15-7.30 (5H).

LC/MS (ESI): 459.2; (calculated ([M+H]⁺): 459.6).

Molecule A21: Product Obtained by Saponification of Molecule A20

Lithium hydroxide (LiOH) (600.49 mg, 25.07 mmol) is added to a solutionof molecule A20 (5.75 g, 12.54 mmol) in a THF/methanol/water mixture(40/40/40 mL) at 0° C., then the mixture is stirred at ambienttemperature for 20 h. After evaporation of the organic solvents under avacuum, the aqueous solution is diluted in water, acidified with anaqueous HCl solution 1 N until the pH is 1. The product is thenextracted with ethyl acetate. The combined organic phases are washedwith a saturated aqueous NaCl solution, dried over Na₂SO₄, filtered andconcentrated at reduced pressure to yield a colorless oil.

Yield: 5.7 g (quantitative).

¹H NMR (CDCl₃, ppm): 0.88 (3H); 1.10-1.30 (16H); 1.50-1.80 (3H);1.67-2.02 (2H); 2.20 (2H); 2.25 (0.4H); 2.60 (0.6H); 2.85-3.10 (2.6H);3.55-3.65 (1.4H); 4.35 (0.6H); 4.55 (0.4H); 4.94 (1H); 6.28 (0.4H); 6.38(0.6H); 7.20-7.30 (5H).

LC/MS (ESI): 445.2; (calculated ([M+H]⁺): 445.6).

Molecule A22: Product Obtained by Reaction Between Boc-ethylenediamineand Molecule A21

By a method similar to the one used for the preparation of molecule A6applied to molecule A21 (5.67 g, 12.75 mmol) and to Boc-ethylenediamine(2.25 g, 14.03 mmol), a colorless oil is obtained after purification bychromatography column on silica gel (dichloromethane, methanol).

Yield: 5.7 g (76%).

¹H NMR (CDCl₃, ppm): 0.88 (3H); 1.25 (16H); 1.43 (9H); 1.58 (2.6H);1.75-1.95 (1.4H); 2.15-2.30 (3H); 2.64 (0.5H); 2.95-3.10 (2.5H);3.20-3.40 (4H); 3.45 (0.5H); 3.55 (0.2H); 3.66 (1H); 4.44 (1H); 4.50(0.2H); 4.60 (0.6H); 4.99 (0.7H); 5.54 (0.5H); 5.95 (0.2H); 6.17 (1H);6.60 (0.5H); 7.07 (0.5H); 7.20-7.40 (5H).

LC/MS (ESI): 587.4; (calculated ([M+H]⁺): 587.8).

Molecule AA9

After a method similar to the one used for the preparation of moleculeAA2 applied to molecule A22 (5.66 g, 9.65 mmol), the residue obtainedafter concentration under a vacuum of the reaction medium is dissolvedin methanol and evaporated at reduced pressure, the operation beingrepeated 4 times to yield a white foam of molecule AA9 in the form of ahydrochloride salt.

Yield: 4.9 g (97%).

¹H NMR (DMSO-d₆, 120° C., ppm): 0.89 (3H); 1.26 (16H); 1.43 (2H); 1.68(0.6H); 1.75-2.00 (3H); 2.05-2.25 (2.4H); 2.82-3.05 (5H); 3.38 (2H);3.50-3.70 (1.4H); 4.25 (0.6H); 4.63 (0.4H); 4.77 (0.6H); 7.25-7.50(5H);7.55-8.20 (4H).

LC/MS (ESI): 487.4; (calculated ([M+H]⁺): 487.7).

EXAMPLE AA10 Molecule AA10 Molecule A23: Product Obtained by theReaction Between Molecule B7 and Boc-ethylenediamine

HOBt (8.94 g, 58.37 mmol) and then Boc-ethylenediamine (112.20 g, 700.00mmol) in solution in DCM (150 mL) are added successively to a solutionof molecule B7 (190.00 g, 583.73 mmol) at 0° C. in DCM (2.9 L). EDC(123.10 g, 642.00 mmol) is then added, then the mixture is stirred for17 h between 0° C. and ambient temperature. The reaction mixture is thenwashed with a saturated aqueous NaHCO₃ (2×1.5 L), an aqueous HClsolution 1 N (2×1.5 L), then a saturated aqueous NaCl solution (1.5 L),dried over Na₂SO₄, filtered and concentrated at reduced pressure. Awhite solid is obtained after recrystallization in acetonitrile.

Yield: 256.50 g (93%).

¹H NMR (CDCl₃, ppm): 0.88 (3H); 1.16-1.38 (20H); 1.44 (9H); 1.56-1.71(2H); 1.78-2.45 (6H); 3.11-3.72 (6H); 4.30 (0.1H); 4.51 (0.9H); 4.87(0.1H); 5.04 (0.9H); 6.87 (0.1H); 7.23 (0.9H).

LC/MS (ESI): 468.0; (calculated ([M+H]⁺): 468.4).

Molecule AA10

According to a method similar to the method used for the preparation ofmolecule AA1 applied to molecule A23 (256.50 g, 548.43 mmol), a whitesolid of molecule AA10 in the form of a hydrochloride salt is obtainedby trituration in pentane (1.6 L) and drying at reduced pressure at 40°C.

Yield: 220.00 g (99%).

¹H NMR (MeOD-d4, ppm): 0.90 (3H); 1.21-1.43 (20H); 1.54-1.66 (2H);1.85-2.28 (4H); 2.39 (2H); 3.00-3.17 (2H); 3.30-3.40 (1H); 3.43-3.71(3H); 4.29 (0.94H); 4.48 (0.06H).

LC/MS (ESI): 368.2; (calculated ([M+H]⁺): 368.3).

EXAMPLE AA11 Molecule AA11 Molecule A24: Product Obtained by theReaction Between Molecule B7 and Boc-1-amino-4,7,10-trioxa-13-tridecaneamine

By a method similar to the one used for the preparation of molecule A23applied to molecule B7 (24.00 g, 73.73 mmol) and toBoc-1-amino-4,7,10-trioxa-13-tridecane amine (28.35 g, 88.48 mmol), anorange oil of molecule A24 is obtained.

Yield: 44.50 g (96%).

¹H NMR (CDCl₃, ppm): 0.87 (3H); 1.08-1.56 (20H); 1.43 (9H); 1.58-1.67(2H); 1.70-2.00 (6H); 2.04-2.41 (4H); 3.16-3.77 (18H); 4.26-4.29 (0.2H);4.50-4.54 (0.8H); 4.68-5.10 (1H); 6.74 (0.2H); 7.19 (0.8H).

LC/MS (ESI): 628.4; (calculated ([M+H]⁺): 628.5).

Molecule AA11

After a method similar to the one used for the preparation for moleculeAA1 applied to molecule A24 (43.40 g, 69.12 mmol), a white solid ofmolecule AA11 in the form of a hydrochloride salt is obtained aftertrituration 3 times in diethyl ether, solubilization of the residue inwater, and lyophilization.

Yield: 38.70 g (98%).

¹H NMR (DMSO, ppm): 0.85 (3H); 1.07-1.38 (20H); 1.41-1.52 (2H);1.55-1.66 (2H); 1.70-2.02 (6H); 2.08-2.30 (2H); 2.78-2.87 (2H);3.00-3.16 (2H); 3.29-3.66 (14H); 4.16-4.22 (0.65 H); 4.25-4.30 (0.35H);7.74 (0.65H); 7.86 (3H); 8.10 (0.35H).

LC/MS (ESI): 528.4; (calculated ([M+H]⁺): 528.4).

EXAMPLE AA12 Molecule AA12 Molecule A25: Product Obtained by theReaction Between Molecule B4 and Boc-ethylenediamine

By a method similar to the one used for the preparation of molecule A23applied to molecule B4 (12.00 g, 40.35 mmol) and to Boc-ethylenediamine(7.76 g, 48.42 mmol), a colorless oil is obtained and used without otherpurification.

Yield: 17.40 g (94%).

¹H NMR (CDCl₃, ppm): 0.86 (3H); 1.11-1.68 (18H); 1.41 (9H); 1.80-2.38(6H); 3.06-3.35 (4H); 3.37-3.49 (1H); 3.51-3.73 (1H); 4.26-4.31 (0.1H);4.45-4.52 (0.9H); 4.91-5.19 (1H); 6.97 (0.1H); 7.23 (0.9H).

LC/MS (ESI): 440.4 (calculated ([M+H]⁺): 440.3).

Molecule AA12

After a method similar to the one used for the preparation of moleculeAA1 applied to molecule A25 (8.85 g, 20.13 mmol), a white solid ofmolecule AA12 is obtained after alkaline washing, concentration atreduced pressure, then recrystallization in acetonitrile.

Yield: 6.53 g (96%).

¹H NMR (DMSO, ppm): 0.85 (3H); 1.07-1.56 (20H); 1.68-2.03 (4H);2.09-2.29 (2H); 2.50-2.58 (2H); 2.96-3.11 (2H); 3.21-3.59 (2H);4.17-4.21 (0.65H); 4.25-4.29 (0.35H); 7.68 (0.65H); 8.00 (0.35H).

LC/MS (ESI): 340.3; (calculated ([M+H]⁺): 340.3).

EXAMPLE AA13 Molecule AA13 Molecule A26: Product Obtained by CouplingBetween Molecule B1 and Boc-ethylenediamine

By a method similar to the one used for the preparation of molecule A23applied to molecule B1 (30.00 g, 111.36 mmol) and to Boc-ethylenediamine(21.41 g, 133.64 mmol), a white solid is obtained afterrecrystallization in acetonitrile.

Yield: 34.90 g (76%).

¹H NMR (CDCl₃, ppm): 0.88 (3H); 1.10-1.70 (14H); 1.43 (9H); 1.80-1.91(1H); 1.92-2.01 (1H); 2.04-2.42 (4H); 3.13-3.70 (6H); 4.27-4.31 (0.15H);4.47-4.53 (0.85H); 4.83 (0.15H); 5.02 (0.85H); 6.85 (0.15H); 7.21(0.85H).

LC/MS (ESI): 412.2; (calculated ([M+H]⁺): 412.3).

Molecule AA13

After a method similar to the one used for the preparation of moleculeAA1 applied to molecule A26 (34.90 g, 84.79 mmol), a white solid ofmolecule AA13 in the form of a hydrochloride salt is obtained aftersolubilization in a mixture of DCM/acetonitrile and concentration atreduced pressure.

Yield: 29.50 g (99%).

¹H NMR (DMSO, ppm): 0.85 (3H); 1.07-1.61 (14H); 1.70-2.06 (4H);2.10-2.35 (2H); 2.76-2.87 (2H); 3.24-3.47 (3.25H); 3.56-3.64 (0.75H);4.13-4.19 (0.75H); 4.31-4.36 (0.25H); 8.05-8.36 (3.75H); 8.50 (0.25H).

LC/MS (ESI): 312.2; (calculated ([M+H]⁺): 312.3).

EXAMPLE AA14 Molecule AA14 Molecule A27: Product Obtained byHydrogenation of Phytol

Platinum oxide (PtO₂, 1.15 g, 6.61 mmol) is added to a solution ofphytol (30.00 g, 101.20 mmol) in THF (450 mL) under argon, and themixture is placed under 1 bar of dihydrogen, then stirred for 4 h atambient temperature. After filtration through celite rinsing with THF, ablack oil of molecule A27 is obtained by concentration at reducedpressure.

Yield: 29.00 g (96%).

¹H NMR (CDCl₃, ppm): 0.84 (6H); 0.86 (6H); 0.89 (3H); 1.00-1.46 (22H);1.46-1.68 (3H); 3.61-3.73 (2H).

Molecule A28: Product Obtained by Oxidation of Molecule A27

Tetrabutylammonium bromide (16.90 g, 52.45 mmol), acetic acid (150 mL,2.62 mol), then KMnO₄ (46.05 g, 291.40 mmol) are added successively insmall fractions, while maintaining the temperature between 16 and 19°C., to a solution of molecule A27 (29.0 g, 97.13 mmol) in a mixture ofdichloroethane/water (485 mL/388 mL). The reaction mixture is thenstirred for 4 h 30 at reflux, cooled to 10° C., then acidified until thepH is 1 with an HCl solution 6 N (20 mL). Na₂SO₃ (53.90 g) is then addedgradually while maintaining the temperature at 10° C., and the mixtureis stirred until the discoloration is complete. Water (200 mL) is added,the phases are separated, and the aqueous phase is extracted with DCM(2×400 mL). The combined organic phases are washed with an aqueoussolution of HCl at 10% (20 mL), water (2×200 mL), a saturated aqueousNaCl solution (200 mL), dried over Na₂SO₄, filtered and concentrated atreduced pressure. A yellow oil of molecule A28 is obtained afterpurification by flash chromatography (eluent: cyclohexane, AcOEt).

Yield: 28.70 g (94%).

¹H NMR (CDCl₃, ppm): 0.84 (6H); 0.86 (6H); 0.97 (3H); 1.00-1.41 (20H);1.52 (1H); 1.96 (1H); 2.14 (1H); 2.35 (1H); 11.31 (1H).

LC/MS (ESI): 311.1 (calculated ([M−H]⁻): 311.3).

Molecule A29: Product Obtained by Coupling Between Molecule A28 andMethyl L-prolinate

By a method similar to the one used for the preparation of molecule A2applied to molecule A28 (18.00 g, 57.59 mmol) and to the hydrochlorideof methyl L-prolinate (14.31 g, 86.39 mmol), a yellow oil of moleculeA29 is obtained after washing of the organic phase with a saturatedaqueous NaHCO₃ solution (2×150 mL), an aqueous solution of HCl at 10%(2×150 mL), a saturated aqueous NaCl solution (2×150 mL), then dryingover Na₂SO₄, filtration and concentration at reduced pressure.

Yield: 23.20 g (95%).

¹H NMR (DMSO-d₆, ppm): 0.78-0.89 (15H); 0.97-1.43 (20H); 1.43-1.56 (1H);1.70-1.96 (4H); 1.96-2.32 (3H); 3.33-3.56 (2H); 3.59 (0.6H); 3.67(2.4H); 4.27 (0.8H); 4.57 (0.2H).

LC/MS (ESI): 424.4 (calculated ([M+H]⁺): 424.4).

Molecule A30: Product Obtained by the Saponification of Molecule A29

By a method similar to the one used for the preparation of molecule A21applied to molecule A29 (21.05 g, 49.68 mmol), a yellow oil of moleculeA30 is obtained.

Yield: 20.40 g (99%).

¹H NMR (DMSO-d₆, ppm): 0.77-0.91 (15H); 0.97-1.43 (20H); 1.43-1.56 (1H);1.67-1.96 (4H); 1.96-2.29 (3H); 3.26-3.56 (2H); 4.20 (0.8H); 4.41(0.2H).

LC/MS (ESI): 410.3 (calculated ([M+H]⁺): 410.4).

Molecule A31: Product Obtained by the Coupling Between Molecule A30 andBoc-1-amino-4,7,10-trioxa-13-tridecane amine

By a method similar to the one used for the preparation of molecule A23applied to molecule A30 (8.95 g, 21.85 mmol) and toBoc-1-amino-4,7,10-trioxa-13-tridecane amine (8.40 g, 26.21 mmol), acolorless oil of molecule A31 is obtained after purification by flashchromatography (eluent: DCM, AcOEt, methanol).

Yield: 10.08 g (65%).

¹H NMR (DMSO-d₆, ppm): 0.78-0.89 (15H); 0.97-1.43 (29H); 1.43-1.55 (1H);1.55-1.66 (4H); 1.71-2.30 (7H); 2.95 (2H); 3.00-3.19 (2H); 3.34-3.58(14H); 4.17-4.29 (1H); 6.30-6.79 (1H); 7.67 (0.65H); 8.00 (0.35H).

LC/MS (ESI): 712.6 (calculated ([M+H]⁺): 712.6).

Molecule AA14

After a method similar to the one used for the preparation of moleculeAA1 applied to molecule A31 (10.08 g, 14.16 mmol), the residue obtainedafter concentration at reduced pressure is solubilized in DCM (200 mL),the organic phase is washed with an aqueous NaOH solution 2 N (2×100mL), dried over Na₂SO₄, filtered and concentrated at reduced pressure. Acolorless oil of molecule AA14 in the form of a neutral amine isobtained.

Yield: 8.23 g (95%).

¹H NMR (DMSO-d₆, ppm): 0.78-0.89 (15H); 0.97-1.43 (20H); 1.43-1.69 (6H);1.69-2.30 (8H); 2.56 (2H); 2.99-3.19 (2H); 3.31-3.58 (14H); 4.15-4.29(1H); 7.70 (0.65H); 8.04 (0.35H).

LC/MS (ESI): 612.5 (calculated ([M+H]⁺): 612.5).

AB: Synthesis of the Co-Polyamino Acids Modified by HydrophobicMolecules in which p=1

Statistical Co-Polyamino Acids of Formula VII or VIIa.

TABLE 1B List of the co-polyamino acids of formula VII or VIIasynthesized according to the invention no co-polyamino acids bearingcarboxylate charges and hydrophobic radicals AB1

AB2

AB3

AB4

AB5

AB6

AB7

AB8

AB9

AB10

AB11

AB12

AB13

AB21

AB22

AB23

AB24

AB25

AB26

AB27

AB28

AB29

AB30

AB31

AB32

Defined Co-Polyamino Acids of Formula VII or VIIb

TABLE 1C list of the co-polyamino acids of formula VII or VIIbsynthesized according to the invention. Example No co-polyamino acidsbearing carboxylate charges and hydrophobic radicals AB14

AB15

AB16

AB17

AB18

AB19

AB20

Co-Polyamino Acids of Formula VII or VIIa EXAMPLE AB1 Co-Polyamino AcidAB1—Sodium poly-L-glutamate Modified by Molecule AA1 and Having a NumberAverage Molecular Weight (Mn) of 2900 g/mol

Co-polyamino acid AB1-1: poly-L-glutamic acid of relative number averagemolecular weight (Mn) 3861 g/mol originating from the polymerization ofγ-benzyl-L-glutamate N-carboxyanhydride initiated by hexylamine.

In a round-bottom flask dried in the oven, γ-benzyl-L-glutamateN-carboxyanhydride (89.9 g, 341 mmol) is placed under a vacuum for 30min, then anhydrous DMF (200 mL) is introduced. The mixture is thenstirred under argon until the dissolution is complete, cooled to 4° C.,then hexylamine (2.05 mL, 15.5 mmol) is introduced rapidly. The mixtureis stirred between 4° C. and ambient temperature for 2 days. Thereaction medium is then heated at 65° C. for 2 h, cooled to ambienttemperature, then poured dropwise into diisopropyl ether (3 L) understirring. The white precipitate is recovered by a filtration, washedwith diisopropyl ether (2×20 mL), then dried under a vacuum at 30° C. toyield a poly(gamma-benzyl-L-glutamic acid) (PBLG).

A solution of hydrobromic acid (HBr) at 33% in acetic acid (240 mL, 1.37mol) is added dropwise to a solution of PBLG (74.8 g) in trifluoroaceticacid (TFA, 340 mL) at 4° C. The mixture is stirred at ambienttemperature for 2 h, then poured dropwise into a 1:1 (v/v) mixture ofdiisopropyl ether and water under stirring (4 L). After 2 h of stirring,the heterogeneous mixture is allowed to stand overnight. The whiteprecipitate is recovered by filtration, washed with a 1:1 (v/v) mixtureof diisopropyl ether and water (340 mL), then with water (340 mL).

The solid obtained is then solubilized in water (1.5 L) by adjusting thepH to 7 by adding an aqueous sodium hydroxide solution 10 N, then anaqueous sodium hydroxide solution 1 N. After solubilization, thetheoretical concentration is adjusted to 20 g/L theoretical by additionof water to obtain a final volume of 2.1 L.

The solution is filtered through a 0.45 μm filter, then purified byultrafiltration against a NaCl solution 0.9%, then water until theconductimetry of the permeate is less than 50 μS/cm. The solution ofco-polyamino acid is then concentrated until a final volume of 1.8 L isobtained.

The aqueous solution is then acidified by adding a hydrochloric acidsolution 37% until a pH of 2 is reached. After 4 h of stirring, theprecipitate obtained is filtered, washed with water (2×340 mL), thendried under a vacuum at 30° C. to yield a poly-L-glutamic acid of numberaverage molecular weight (Mn) 3861 g/mol with respect to apolyoxyethylene standard (PEG).

Co-Polyamino Acid AB1

The co-polyamino acid AB1-1 (10.0 g) is solubilized in DMF (700 mL) at30° C., then cooled to 0° C. Molecule AA1 in the form of a hydrochloridesalt (1.64 g, 3.8 mmol) is suspended in DMF (23 mL), and triethylamine(0.39 g, 3.8 mmol) is then added, and the mixture is heated slightlyunder stirring until the dissolution is complete. N-methylmorpholine(NMM, 7.6 g, 75 mmol) in DMF (14 mL) and ethyl chloroformate (ECF, 8.2g, 75 mmol) are added to the solution of co-polyamino acid at 0° C.After 10 min at 0° C., the solution containing molecule AA1 is added andthe mixture is maintained at 30° C. for 2 h. The reaction mixture ispoured dropwise into 5.5 L of water containing NaCl at 15 wt % and HCl(pH 2), and then allowed to stand overnight. The precipitate iscollected by filtration and dried under a vacuum for approximately 30min. The white solid obtained is dissolved in water (500 mL), and the pHis adjusted to 7 by slow addition of an aqueous NaOH solution 1 N. Afterfiltration through a 0.45 μm filter, the clear solution obtained ispurified by ultrafiltration against a solution of NaCl 0.9%, then water,until the conductimetry of the permeate is less than 50 μS/cm. Afterremoval, the solution is filtered through a 0.2 μm filter and stored at2-8° C.

-   Dry extract: 24.9 mg/g.-   An average degree of polymerization (DP) of 23 is estimated by ¹H    NMR in D₂O by comparing the integration of the signals from the    grafted hydrophobe with the integration of the signals from the main    chain.-   Based on ¹H NMR: i=0.05.-   The calculated average molecular weight of co-polyamino acid AB1 is    calculated based on the molecular weights of the radicals R₁ and R₂,    the aspartate and/or glutamate residues (including an amide bond),    the hydrophobic radical, DS and DP.-   The calculated average molecular weight of co-polyamino acid AB1 is    3945 g/mol.-   HPLC-aqueous SEC (calibrant PEG): Mn=2900 g/mol.

EXAMPLE AB2 Co-Polyamino Acid AB2—Sodium poly-L-glutamate Modified byMolecule AA1 and Having a Number Average Molecular Weight (Mn) of 3700g/mol

By a method similar to the one used for the preparation of co-polyaminoacid AB1 applied to the hydrochloride salt of molecule AA1 (1.64 g, 3.8mmol) and to a poly-L-glutamic acid of relative Mn 5200 g/mol (10.0 g)obtained by a method similar to the one used for the preparation ofco-polyamino acid AB1-1, a sodium poly-L-glutamate modified by moleculeAA1 is obtained.

-   Dry extract: 14.1 mg/g.-   DP (estimated based on ¹H NMR): 35.-   Based on ¹H NMR: i=0.05.-   The calculated average molecular weight of co-polyamino acid AB2 is    5972 g/mol.-   HPLC-aqueous SEC (calibrant PEG): Mn=3700 g/mol.

EXAMPLE AB3 Co-Polyamino Acid AB3—Sodium poly-L-glutamate Modified byMolecule AA1 and Having a Number Average Molecular Weight (Mn) of 4900g/mol

By a method similar to the one used for the preparation of co-polyaminoacid AB1 applied to the hydrochloride salt of molecule AA1 (3.30 g, 7.6mmol) and to a poly-L-glutamic acid of relative number average weight(Mn) 5200 g/mol (10.0 g) obtained by a method similar to the one usedfor the preparation of co-polyamino acid AB1-1, a sodiumpoly-L-glutamate modified by molecule AA1 is obtained.

-   Dry extract: 23.4 mg/g.-   DP (estimated based on ¹H NMR): 35.-   The calculated average molecular weight of co-polyamino acid AB3 is    6594 g/mol.-   Based on ¹H NMR: i=0.10.-   HPLC-aqueous SEC (calibrant PEG): Mn=4900 g/mol.

EXAMPLE AB4 Co-Polyamino Acid AB4—Sodium poly-L-glutamate Modified byMolecule AA2 and Having a Number Average Molecular Weight (Mn) of 1800g/mol

By a method similar to the one used for the preparation of co-polyaminoacid AB1 applied to the hydrochloride salt of molecule AA2 (1.09 g, 2.4mmol) and to a poly-L-glutamic acid of average weight Mn=5600 g/mol (6.3g) obtained by a method similar to the one used for the preparation ofco-polyamino acid AB1-1, but with a step of deprotection of the benzylesters using trimethylsilane iodide according to the protocol describedin the publication Subramanian G. et al., J. Am. Chem. Soc. 2000, 122,26-34, a sodium poly-L-glutamate modified by molecule AA2 is obtained.

-   Dry extract: 21.5 mg/g.-   DP (estimated based on ¹H NMR): 35.-   Based on ¹H NMR: i=0.052.-   The calculated average molecular weight of co-polyamino acid AB4 is    6022 g/mol.-   HPLC-aqueous SEC (calibrant PEG): Mn=1800 g/mol.

EXAMPLE AB5 Co-Polyamino Acid AB5—Sodium poly-L-glutamate Modified byMolecule AA6 and Having a Number Average Molecular Weight (Mn) of 2600g/mol

By a method similar to the one used for the preparation of co-polyaminoacid AB1 applied to the hydrochloride salt of molecule AA6 (2.06 g, 3.8mmol) and to a poly-L-glutamic acid (9.8 g) obtained by a method similarto the one used for the preparation of co-polyamino acid AB1-1, a sodiumpoly-L-glutamate modified by molecule AA6 is obtained.

-   Dry extract: 20.9 mg/g.-   DP (estimated based on 1H NMR): 23.-   Based on 1H NMR: i=0.05.-   The calculated average molecular weight of co-polyamino acid AB5 is    4079 g/mol.-   HPLC-aqueous SEC (calibrant PEG): Mn=2600 g/mol.

EXAMPLE AB6 Co-Polyamino Acid AB6—Sodium poly-L-glutamate Modified byMolecule AA7 and Having a Number Average Molecular Weight (Mn) of 4000g/mol

A poly-L-glutamic acid of average weight Mn=3500 g/mol and degree ofpolymerization 22 (10.0 g) obtained by a method similar to the one usedfor the preparation of co-polyamino acid AB1-1 is solubilized in DMF(420 mL) at 30-40° C., then maintained at this temperature. In parallel,the hydrochloride salt of molecule AA7 (1.47 g, 2.3 mmol) is suspendedin DMF (12 mL), and triethylamine (0.23 g, 2.3 mmol) is added, then themixture is heated slightly under stirring until the dissolution iscomplete. NMM (7.6 g, 75 mmol), the solution of AA7 and then the N-oxideof 2-hydroxypyridine (HOPO, 0.84 g, 7.5 mmol) are added successively tothe solution of co-polyamino acid in DMF. The reaction medium is thencooled to 0° C., then EDC (1.44 g, 7.5 mmol) is added, and the medium isbrought again to ambient temperature in 2 h. The reaction medium isfiltered through a 0.2 mm woven filter and poured dropwise into 3.5 L ofwater containing NaCl at 15 wt % and HCl (pH 2) under stirring. At theend of the addition, the pH is readjusted to 2 with an HCl solution 37%,and the suspension is allowed to stand overnight. The precipitate iscollected by filtration, then rinsed with 100 mL of water. The whitesolid obtained is solubilized in 500 mL of water by slow addition of anaqueous solution of NaOH 1 N until the pH is 7 under stirring, then thesolution is filtered through a 0.45 μm filter. The clear solutionobtained is purified by ultrafiltration against a solution of NaCl 0.9%,then water, until the conductimetry of the permeate is less than 50μS/cm. The solution is filtered through a 0.2 μm filter and stored at2-8° C.

-   Dry extract: 21.6 mg/g.-   DP (estimated based on 1H NMR): 20.-   Based on 1H NMR: i=0.025.-   The calculated average molecular weight of co-polyamino acid AB6 is    3369 g/mol.-   HPLC-aqueous SEC (calibrant PEG): Mn=4000 g/mol.

EXAMPLE AB7 Co-Polyamino Acid AB7—Sodium poly-L-glutamate Capped at Oneof its Ends by an Acetyl Group and Modified by Molecule AA7 and Having aNumber Average Molecular Weight (Mn) of 3300 g/mol

Co-polyamino acid AB7-1: poly-L-glutamic acid of relative number averagemolecular weight (Mn) 3600 g/mol and DP 21 originating from thepolymerization of γ-benzyl-L-glutamate N-carboxyanhydride, initiated byhexylamine and capped at one of its ends by an acetyl group.

γ-Benzyl-L-glutamate N-carboxyanhydride (Glu(OBn)-NCA, 100.0 g, 380mmol) is placed for 30 min under a vacuum in a round-bottom flask driedin the oven, then anhydrous DMS (225 mL) is introduced. The mixture isthen stirred under argon until the dissolution is complete, cooled to 4°C., then hexylamine (1.78 g, 17 mmol) is introduced rapidly. The mixtureis stirred between 4° C. and ambient temperature for 2 days, thenprecipitated in diisopropyl ether (3.4 L). The precipitate is recoveredby filtration, washed two times with diisopropyl ether (225 mL), thendried to yield a white solid which is dissolved in 450 mL of THF. DIPEA(31 mL, 176 mmol) and then acetic anhydride (17 mL, 176 mmol) are addedsuccessively to this solution. After stirring overnight at ambienttemperature, the solution is poured slowly into diisopropyl ether (3 L)under stirring. After 1 h of stirring, the precipitate is filtered,washed two times with diisopropyl ether (250 mL), then dried under avacuum at 30° C. to yield a poly(gamma-benzyl-L-glutamic acid) capped atone of its ends by an acetyl group.

A solution of hydrobromic acid (HBr) at 33% in acetic acid (235 mL) isadded dropwise to a solution of the above co-polyamino acid (72 g) intrifluoroacetic acid (TFA, 335 mL) at 4° C. The mixture is stirred atambient temperature for 3 h 30, then poured dropwise into a 1:1 (v/v)mixture of diisopropyl ether and water under stirring (4 L). After 2 hof stirring, the heterogeneous mixture is allowed to stand overnight.The white precipitate is recovered by filtration, washed with a 1:1(v/v) mixture of diisopropyl ether and water (340 mL), then with water(340 mL).

The solid obtained is then solubilized in water (1.5 L) by adjusting thepH to 7 by addition of an aqueous solution of sodium hydroxide 10 N,then an aqueous solution of sodium hydroxide 1 N. After solubilization,the solution is diluted by addition of water to obtain a final volume of2.1 L. The solution is filtered through a 0.45 μm filter, then purifiedby ultrafiltration against a solution of NaCl 0.9%, then water until theconductimetry of the permeate is less than 50 μS/cm. The solution ofco-polyamino acid is then concentrated until a final volume of 1.8 L isobtained.

The aqueous solution is then acidified by addition of a hydrochloricacid solution 37% until a pH of 2 is reached. After 4 h of stirring, theprecipitate obtained is filtered, washed with water (330 mL), then driedunder a vacuum at 30° C. to yield a poly-L-glutamic acid of numberaverage molecular weight (Mn) 3600 g/mol with respect to apolyoxyethylene standard (PEG) and of average degree of polymerization21.

Co-Polyamino Acid AB7:

By a method similar to the one used for the preparation of co-polyaminoacid AB6 applied to the hydrochloride salt of molecule AA7 (1.43 g, 2.2mmol) and to co-polyamino acid AB7-1 (10.0 g), a sodium poly-L-glutamateacid modified by molecule AA7 is obtained.

-   Dry extract: 24.3 mg/g.-   DP (estimated based on 1H NMR): 21.-   Based on 1H NMR: i=0.03.-   The calculated average molecular weight of co-polyamino acid AB7 is    3677 g/mol.-   HPLC-aqueous SEC (calibrant PEG): Mn=3300 g/mol.

EXAMPLE AB8 Co-Polyamino Acid AB8—Sodium poly-L-glutamate Modified byMolecule AA7 and Having a Number Average Molecular Weight (Mn) of 3600g/mol

Co-polyamino acid AB8-1: poly-L-glutamic acid of number averagemolecular weight (Mn) 3800 g/mol and degree of polymerization 24originating from the polymerization of γ-methyl-L-glutamateN-carboxyanhydride initiated by ammonia.

By a method similar to the one described in the patent applicationFR-A-2 801 226 applied to γ-methyl-L-glutamic acid N-carboxyanhydride(25.0 g, 133.6 mmol) and to an ammonia solution 0.5 N in dioxane (12.1mL, 6.05 mmol), a poly-L-glutamic acid is obtained.

Co-Polyamino Acid AB8:

By a method similar to the one used for the preparation of co-polyaminoacid AB6 applied to the hydrochloride salt of molecule AA7 (2.1 g, 3.24mmol) and to co-polyamino acid AB8-1 (14.3 g), a sodium poly-L-glutamatemodified by molecule AA7 is obtained.

-   Dry extract: 25.2 mg/g.-   DP (estimated based on 1H NMR): 24.-   Based on ¹H NMR: i=0.03.-   The calculated average molecular weight of co-polyamino acid AB8 is    4099 g/mol.-   HPLC-aqueous SEC (calibrant PEG): Mn=3600 g/mol.

EXAMPLE AB9 Co-Polyamino Acid AB9—Sodium poly-L-glutamate Modified byMolecule AA3 and Having a Number Average Molecular Weight (Mn) of 3200g/mol

By a method similar to the one used for the preparation of co-polyaminoacid AB1 applied to the hydrochloride salt of molecule AA3 and to apoly-L-glutamic acid obtained by a method similar to the one used forthe preparation of co-polyamino acid AB1-1, a sodium poly-L-glutamatemodified by molecule AA3 is obtained.

-   Dry extract: 14.7 mg/g.-   DP (estimated based on ¹H NMR): 30.-   Based on ¹H NMR: i=0.12.-   The calculated average molecular weight of co-polyamino acid AB9 is    6192 g/mol.-   HPLC-aqueous SEC (calibrant PEG): Mn=3200 g/mol.

EXAMPLE AB10 Co-polyamino acid AB10—Sodium poly-L-glutamate Modified byMolecule AA4 and Having a Number Average Molecular Weight (Mn) of 2600g/mol

By a method similar to the one used for the preparation of co-polyaminoacid AB7 applied to the hydrochloride salt of molecule AA4 and to apoly-L-glutamic acid obtained by a method similar to the one used forthe preparation of co-polyamino acid AB1-1, a sodium poly-L-glutamatemodified by molecule AA4 is obtained.

-   Dry extract: 18.3 mg/g.-   DP (estimated based on ¹H NMR): 25.-   Based on ¹H NMR: i=0.08.-   The calculated average molecular weight of co-polyamino acid AB10 is    4870 g/mol.-   HPLC-aqueous SEC (calibrant PEG): Mn=2600 g/mol.

EXAMPLE AB11 Co-Polyamino Acid AB11—Sodium poly-L-glutamate Modified byMolecule AA5 and Having a Number Average Molecular Weight (Mn) of 2700g/mol

By a method similar to the one used for the preparation of co-polyaminoacid AB6 applied to the hydrochloride salt of molecule AA5 and to apoly-L-glutamic acid obtained by a method similar to the one used forthe preparation of co-polyamino acid AB1-1, a sodium poly-L-glutamatemodified by molecule AA5 is obtained.

-   Dry extract: 20.2 mg/g.-   DP (estimated based on ¹H NMR): 23.-   Based on ¹H NMR: i=0.05.-   The calculated average molecular weight of co-polyamino acid AB11 is    4072 g/mol.-   HPLC-aqueous SEC (calibrant PEG): Mn=2700 g/mol.

EXAMPLE AB12 Co-Polyamino Acid AB12—Sodium poly-L-glutamate Modified byMolecule AA8 and Having a Number Average Molecular Weight (Mn) of 3000g/mol

By a method similar to the one used for the preparation of co-polyaminoacid AB1 applied to the hydrochloride salt of molecule AA8 and to apoly-L-glutamic acid obtained by a method similar to the one used forthe preparation of co-polyamino acid AB1-1, a sodium poly-L-glutamatemodified by molecule AA8 is obtained.

-   Dry extract: 19.5 mg/g.-   DP (estimated based on ¹H NMR): 26.-   Based on ¹H NMR: i=0.04.-   The calculated average molecular weight of co-polyamino acid AB12 is    4477 g/mol.-   HPLC-aqueous SEC (calibrant PEG): Mn=3000 g/mol.

EXAMPLE AB13 Co-Polyamino Acid AB13—Sodium poly-L-glutamate Modified byMolecule AA9 and Having a Number Average Molecular Weight (Nm) of 3300g/mol

By a method similar to the one used for the preparation of co-polyaminoacid AB6 applied to the hydrochloride salt of molecule AA9 and to apoly-L-glutamic acid obtained by a method similar to the one used forthe preparation of co-polyamino acid AB1-1 using isoamylamine asinitiator instead of hexylamine, a sodium poly-L-glutamate modified bymolecule AA9 is obtained.

-   Dry extract: 22.3 mg/g.-   DP (estimated based on ¹H NMR): 35.-   Based on ¹H NMR: i=0.12.-   The calculated average molecular weight of co-polyamino acid AB13 is    7226 g/mol.-   HPLC-aqueous SEC (calibrant PEG): Mn=3300 g/mol.

EXAMPLE AB21 Co-Polyamino Acid AB21—Sodium poly-L-glutamate Modified byMolecule AA7 and Having a Number Average Molecular Weight (Mn) of 3400g/mol

By a method similar to the one used for the preparation of co-polyaminoacid AB6 applied to the hydrochloride salt of molecule AA7 (2.44 g, 2.4mmol) and to a poly-L-glutamic acid (10 g) obtained by a method similarto the one used for the preparation of co-polyamino acid AB1-1, a sodiumpoly-L-glutamate modified by molecule AA7 is obtained.

-   Dry extract: 22.7 mg/g.-   DP (estimated based on ¹H NMR): 22.-   Based on ¹H NMR: i=0.056.-   The calculated average molecular weight of co-polyamino acid AB21 is    4090 g/mol.-   HPLC-aqueous SEC (calibrant PEG): Mn=3400 g/mol.

EXAMPLE AB22 Co-Polyamino Acid AB22—Sodium poly-L-glutamate Capped atOne of its Ends by an Acetyl Group and Modified by Molecule AA10 andHaving a Number Average Molecular Weight (Mn) of 4000 g/mol

The hydrochloride salt of molecule AA10 (4.56 g, 11.29 mmol) isdissolved in chloroform (60 mL) and triethylamine (1.14 g, 11.29 mmol)is added. To a solution of co-polyamino acid (10.0 g, 75.3 mmol)obtained by a method similar to the one used for the preparation ofco-polyamino acid B7-1 in DMF (420 mL), NMM (7.6 g, 75.26 mmol), thenHOPO (2.51 g, 22.58 mmol) are added successively. The reaction medium isthen cooled to 0° C., then EDC (4.33 g, 22.58 mmol) is added, the mediumis stirred for 1 h at 0° C., then the solution of molecule AA10 isadded. The reaction mixture is stirred for 2 h between 0° C. and ambienttemperature. The reaction medium is filtered through a 0.2 mm wovenfilter and poured dropwise into 3.95 L of water containing NaCl at 15 wt% and HCl (pH 2) under stirring. At the end of the addition, the pH isreadjusted to 2 with a solution of HCl 37% and the suspension is allowedto stand overnight. The precipitate is collected by a filtration, thensolubilized in 780 mL of water by slow addition of an aqueous NaOHsolution 1 N until the pH is 7 under stirring. After filtration througha 0.45 μm filter, the solution is diluted by addition of water, thenacetone is added to obtain a solution containing 30 wt % of acetone.This solution is filtered through an activated charcoal filter, then theacetone is distilled (40° C., 100 mbar). After filtration through a 0.45μm filter, the product is purified by ultrafiltration against an aqueoussolution of NaCl at 0.9%, a carbonate buffer solution (150 mM), anaqueous solution of NaCl at 0.9%, a phosphate buffer solution (150 mM),an aqueous solution of NaCl at 0.9%, then water until the conductimetryof the permeate is less than 50 μS/cm. The solution is thenconcentrated, filtered through a 0.2 μm filter and stored at 2-8° C.

-   Dry extract: 19.7 mg/g.-   DP (estimated based on ¹H NMR): 38.-   Based on ¹H NMR: i=0.16.-   The calculated average molecular weight of co-polyamino acid AB22 is    7877 g/mol.-   HPLC-organic SEC (calibrant PEG): Mn=4000 g/mol.

EXAMPLE AB23 Co-Polyamino Acid AB23—Sodium poly-L-glutamate and Modifiedby Molecule AA10 and Having a Number Average Molecular Weight (Mn) of7600 g/mol

Co-polyamino acid AB23-1: poly-L-glutamic acid originating from thepolymerization of γ-benzyl-L-glutamate N-carboxyanhydride initiated byhexylamine and capped at one of its ends by a pyroglutamate group.

A poly-L-glutamic acid (20.0 g) obtained by a method similar to the oneused for the preparation of co-polyamino acid AB1-1 is solubilized inDMF at 80° C., then maintained at this temperature. After 24 h, thereaction medium is poured into a solution of NaCl at 15% and at pH 2.After 4 h, the white solid is collected by filtration, rinsed withwater, then dried under a vacuum at 30° C.

Co-Polyamino Amide AB23

By a method similar to the one used for the preparation of co-polyaminoacid AB22 applied to the hydrochloride salt of molecule AA10 (2.742 g,6.79 mmol) and to co-polyamino acid AB23-1 (9.0 g), a sodiumpoly-L-glutamate acid modified by molecule AA10 is obtained.

-   Dry extract: 21.9 mg/g.-   DP (estimated based on ¹H NMR): 60.-   Based on ¹H NMR: i=0.1.-   The calculated average molecular weight of co-polyamino acid AB23 is    11,034 g/mol.-   HPLC-organic SEC (calibrant PEG): Mn=7600 g/mol.

EXAMPLE AB24 Co-Polyamino Acid AB24—Sodium poly-L-glutamate and Modifiedby Molecule AA10 and Having a Number Average Molecular Weight (Mn) of4300 g/mol

By a method similar to the one used for the preparation of co-polyaminoacid AB23 applied to the hydrochloride salt of molecule AA10 and to apoly-L-glutamic acid obtained by a method similar to the one used forthe preparation of co-polyamino acid AB23-1, a sodium poly-L-glutamatemodified by molecule AA10 is obtained.

-   Dry extract: 22.9 mg/g.-   DP (estimated based on ¹H NMR): 39.-   Based on ¹H NMR: i=0.15.-   The calculated average molecular weight of co-polyamino acid AB24 is    7870 g/mol.-   HPLC-organic SEC (calibrant PEG): Mn=4300 g/mol.

EXAMPLE AB25 Co-Polyamino Acid AB25—Sodium poly-L-glutamate and Modifiedby Molecule AA10 and Having a Number Average Molecular Weight (Mn) of4200 g/mol

By a method similar to the one used for the preparation of co-polyaminoacid AB23 applied to the hydrochloride salt of molecule AA10 and to apoly-L-glutamic acid obtained by a method similar to the one used forthe preparation of co-polyamino acid AB23-1, a sodium poly-L-glutamatemodified by molecule AA10 is obtained.

-   Dry extract: 25.9 mg/g.-   DP (estimated based on ¹H NMR): 39.-   Based on ¹H NMR: i=0.2.-   The calculated average molecular weight of co-polyamino acid AB25 is    8509 g/mol.-   HPLC-organic SEC (calibrant PEG): Mn=4200 g/mol.

EXAMPLE AB26 Co-Polyamino Acid AB26—Sodium poly-L-glutamate and Modifiedby Molecule AA10 and Having a Number Average Molecular Weight (Mn) of2700 g/mol

By a method similar to the one used for the preparation of co-polyaminoacid AB23 applied to the hydrochloride salt of molecule AA10 and to apoly-L-glutamic acid obtained by a method similar to the one used forthe preparation of co-polyamino acid AB23-1, a sodium poly-L-glutamatemodified by molecule AA10 is obtained.

-   Dry extract: 23.9 mg/g.-   DP (estimated based on ¹H NMR): 22.-   Based on ¹H NMR: i=0.21.-   The calculated average molecular weight of co-polyamino acid AB26 is    4899 g/mol.-   HPLC-organic SEC (calibrant PEG): Mn=2700 g/mol.

EXAMPLE AB27 Co-Polyamino Acid AB27—Sodium poly-L-glutamate and Modifiedby Molecule AA11 and Having a Number Average Molecular Weight (Mn) of4500 g/mol

By a method similar to the one used for the preparation of co-polyaminoacid AB23 applied to the hydrochloride salt of molecule AA11 and to apoly-L-glutamic acid obtained by a method similar to the one used forthe preparation of co-polyamino acid AB23-1, a sodium poly-L-glutamatemodified by molecule AA11 is obtained.

-   Dry extract: 26.8 mg/g.-   DP (estimated based on ¹H NMR): 39.-   Based on ¹H NMR: i=0.15.-   The calculated average molecular weight of co-polyamino acid AB27 is    8808 g/mol.-   HPLC-organic SEC (calibrant PEG): Mn=4500 g/mol.

EXAMPLE AB28 Co-Polyamino Acid AB28—Sodium poly-L-glutamate and Modifiedby Molecule AA12 and Having a Number Average Molecular Weight (Mn) of4000 g/mol

By a method similar to the one used for the preparation of co-polyaminoacid AB23 applied to the hydrochloride salt of molecule AA12 and to apoly-L-glutamic acid obtained by a method similar to the one used forthe preparation of co-polyamino acid AB23-1, a sodium poly-L-glutamatemodified by molecule AA12 is obtained.

-   Dry extract: 22.9 mg/g.-   DP (estimated based on ¹H NMR): 39.-   Based on ¹H NMR: i=0.15.-   The calculated average molecular weight of co-polyamino acid AB28 is    7706 g/mol.-   HPLC-organic SEC (calibrant PEG): Mn=4000 g/mol.

EXAMPLE AB29 Co-Polyamino Acid AB29—Sodium poly-L-glutamate and Modifiedby Molecule AA13 and Having a Number Average Molecular Weight (Mn) of4000 g/mol

Co-polyamino acid B29-1: poly-L-glutamic acid originating from thepolymerization of γ-benzyl-L-glutamate N-carboxyanhydride initiated byhexylamine. In a double-jacket reactor, γ-benzyl-L-glutamateN-carboxyanhydride (500 g, 1.90 mol) is solubilized in anhydrous DMF(1100 mL). The mixture is then stirred until the dissolution iscomplete, cooled to 0° C., then hexylamine (6.27 mL, 47.5 mmol) isintroduced rapidly. The mixture is stirred at 0° C. for 5 h, between 0°C. and 20° C. for 7 h, then at 20° C. for 7 h. The reaction mixture isthen heated at 65 ° C. for 2 h, cooled to 55° C., and methanol (3300 mL)is introduced in 1 h 30. The reaction mixture is then cooled to 0° C.and left under stirring for 18 h. The white precipitate is recovered byfiltration, washed with diisopropyl ether (2×800 mL), then dried atreduced pressure at 30° C. to yield a poly(gamma-benzyl-L-glutamic acid)(PBLG).

To a solution of PBLG (180 g) in N,N-dimethylacetamide (DMAc, 450 mL),Pd/Al₂O₃ (36 g) is added under an argon atmosphere. The mixture isplaced under a hydrogen atmosphere (10 bar) and stirred at 60° C. for 24h. After cooling at ambient temperature and filtration of the catalystthrough a sintered filter P4, then through a 0.2 μm Omnipore membranehydrophilic PTFE, a solution of water at pH 2 (2700 mL) is addeddropwise to the solution of DMAc, over a period of 45 min and understirring. After 18 h under stirring, the white precipitate is recoveredby filtration, washed with water, then dried at reduced pressure at 30°C.

Co-Polyamino Acid AB29

By a method similar to the one used for the preparation of co-polyaminoacid AB23 applied to the hydrochloride salt of molecule AA13 and toco-polyamino acid AB29-1, a sodium poly-L-glutamate modified by moleculeAA13 is obtained.

-   Dry extract: 16.1 mg/g.-   DP (estimated based on ¹H NMR): 40.-   Based on ¹H NMR: i=0.15.-   The calculated average molecular weight of co-polyamino acid AB29 is    7734 g/mol.-   HPLC-organic SEC (calibrant PEG): Mn=4000 g/mol.

EXAMPLE AB30 Co-Polyamino Acid AB30—Sodium poly-L-glutamate and Modifiedby Molecule AA10 and Having a Number Average Molecular Weight (Mn) of4300 g/mol

By a method similar to the one used for the preparation of co-polyaminoacid AB29 applied to the hydrochloride salt of molecule AA10 and to apoly-L-glutamic acid obtained by a method similar to the one used forthe preparation of co-polyamino acid AB29-1 using a molecule AA10 asinitiator instead of hexylamine, a sodium poly-L-glutamate modified bymolecule AA10 is obtained.

-   Dry extract: 29.2 mg/g.-   DP (estimated based on ¹H NMR): 40.-   Based on ¹H NMR: i=0.125.-   The calculated average molecular weight of co-polyamino acid AB30 is    7682 g/mol.-   HPLC-organic SEC (calibrant PEG): Mn=4300 g/mol.

EXAMPLE AB31 Co-Polyamino Acid AB30—Sodium poly-L-glutamate and Modifiedby Molecule AA10 and Having a Number Average Molecular Weight (Mn) of6300 g/mol

By a method similar to the one used for the preparation of co-polyaminoacid AB29 applied to the hydrochloride salt of molecule AA10 and to apoly-L-glutamic acid obtained by a method similar to the one used forthe preparation of co-polyamino acid AB29-1 using molecule AA10 asinitiator instead of hexylamine, a sodium poly-L-glutamate modified bymolecule AA10 is obtained.

-   Dry extract: 23.1 mg/g.-   DP (estimated based on ¹H NMR): 40.-   Based on ¹H NMR: i=0.175.-   The calculated average molecular weight of co-polyamino acid AB31 is    8337 g/mol.-   HPLC-organic SEC (calibrant PEG): Mn=6300 g/mol.

EXAMPLE AB32 Co-Polyamino acid AB32—Sodium poly-L-glutamate and Modifiedby Molecule AA14 and Having a Number Average Molecular Weight (Mn) of4700 g/mol

By a method similar to the one used for the preparation of co-polyaminoacid AB29applied to molecule AA14 and to poly-L-glutamic acid AB29-1, asodium poly-L-glutamate modified by molecule AA14 is obtained.

-   Dry extract: 13.5 mg/g.-   DP (estimated based on ¹H NMR): 40.-   Based on ¹H NMR: i=0.109.-   The calculated average molecular weight of co-polyamino acid AB32 is    8599 g/mol.-   HPLC-organic SEC (calibrant PEG): Mn=4700 g/mol.

Co-Polyamino Acids Defined by Formula VII or VIIb EXAMPLE AB14Co-Polyamino Acid AB14—Sodium poly-L-glutamate Modified at One of itsEnds by Molecule AA1 and Having a Number Average Molecular Weight (Mn)of 3400 g/mol

The hydrochloride salt of molecule AA1 (2.03 g, 4.70 mmol), chloroform(5 mL), molecular mesh 4 Å (1.3 g) as well as the ion exchange resinAmberlite IRN 150 (1.3 g) are introduced successively into a suitablecontainer. After 1 h of stirring on rollers, the medium is filtered andthe resin is rinsed with chloroform. The mixture is evaporated, thenco-evaporated with toluene. The residue is solubilized in anhydrous DMF(30 mL) to be used directly in the polymerization reaction.

γ-Benzyl-L-glutamate N-carboxyanhydride (25.59 g, 97.2 mmol) is placedunder a vacuum for 30 min in a round-bottom flask dried in the oven,then anhydrous DMF (140 mL) is introduced. The mixture is stirred underargon until the solubilization is complete, cooled at 4° C., then thesolution of molecule AA1 prepared as described above is introducedrapidly. The mixture is stirred between 4° C. and ambient temperaturefor 2 days, then heated at 65° C. for 2 h. The reaction mixture is thencooled to ambient temperature, then poured dropwise into diisopropylether (1.7 L) under stirring. The white precipitate is recovered byfiltration, washed two times with diisopropyl ether (140 mL), then driedunder a vacuum at 30° C. to obtain a white solid. The solid is dilutedin TFA (160 mL), and a solution of hydrobromic acid (HBr) at 33% inacetic acid (62 mL, 354 mmol) is then added dropwise and at 0° C. Thesolution is stirred for 2 h at ambient temperature, then poured dropwiseinto a 1:1 (v/v) mixture of diisopropyl ether/water and under stirring(1.9 L). After 2 h of stirring, the heterogeneous mixture is allowed tostand overnight. The white precipitate is recovered by filtration,washed successively with a 1:1 (v/v) mixture of diisopropyl ether andwater (280 mL), then with water (140 mL). The solid obtained issolubilized in water (530 mL) by adjusting the pH to 7 by addition of anaqueous sodium hydroxide solution 10 N, then an aqueous sodium hydroxidesolution 1 N. After solubilization, the theoretical concentration isadjusted to 20 g/L theoretical by addition of water to obtain a finalvolume of 800 mL. The mixture is filtered through a 0.45 μm filter, thenpurified by ultrafiltration against a solution of NaCl 0.9%, then wateruntil the conductimetry of the permeate is less than 50 μS/cm. Thesolution of co-polyamino acid is then concentrated to approximately 30g/L theoretical and the pH is adjusted to 7.0. The aqueous solution isfiltered through a 0.2 μm filter and stored at 4° C.

-   Dry extract: 24.1 mg/g.-   DP (estimated by ¹H NMR)=25, thus i=0.04.-   The calculated average molecular weight of co-polyamino acid AB14 is    3378 g/mol.-   HPLC-aqueous SEC (calibrant PEG): Mn=3400 g/mol.

EXAMPLE AB15 Co-Polyamino Acid AB15—Sodium poly-L-glutamate Modified atOne of its Ends by Molecule AA6 and Having a Number Average MolecularWeight (Mn) 4100 g/mol

By a method similar to the one used for the preparation of co-polyaminoacid AB14 applied to the hydrochloride salt of molecule AA6 (2.16 g,3.94 mmol) and to 25.58 g (97.2 mmol) of γ-benzyl-L-glutamateN-carboxyanhydride, a sodium poly-L-glutamate modified at one of itsends by molecule AA6 is obtained.

-   Dry extract: 45.5 mg/g.-   DP (estimated by ¹H NMR)=30, thus i=0.033.-   The calculated average molecular weight of co-polyamino acid AB15 is    5005 g/mol.-   HPLC-aqueous SEC (calibrant PEG): Mn=4100 g/mol.

EXAMPLE AB16 Co-Polyamino Acid AB16—Sodium poly-L-glutamate Modified atOne of its Ends by Molecule AA6 and Having a Number Average MolecularWeight (Mn) of 6500 g/mol

By a method similar to the one used for the preparation of co-polyaminoacid AB14 applied to the hydrochloride salt of molecule AA6 (2.39 g,4.36 mmol) and to 50.0 g (189.9 mmol) of γ-benzyl-L-glutamateN-carboxyanhydride, a sodium poly-L-glutamate modified at one of itsends by molecule AA6 is obtained.

-   Dry extract: 28.5 mg/g.-   DP (estimated by ¹H NMR)=48, thus i=0.021.-   The calculated average molecular weight of co-polyamino acid AB16 is    7725 g/mol.-   HPLC-aqueous SEC (calibrant PEG): Mn=6500 g/mol.

EXAMPLE AB17 Co-Polyamino Acid AB17—Sodium poly-L-glutamate Modified atOne of its Ends by Molecule AA7 and Having a Number Average MolecularWeight (Mn) of 3500 g/mol

By a method similar to the one used for the preparation of co-polyaminoacid AB14 applied to the hydrochloride salt of molecule AA7 (2.80 g,4.32 mmol) and to 25.0 g (94.9 mmol) of γ-benzyl-L-glutamateN-carboxyanhydride, a sodium poly-L-glutamate modified at one of itsends by molecule AA7 is obtained

-   Dry extract: 25.2 mg/g.-   DP (estimated by ¹H NMR)=26, thus i=0.038.-   The calculated average molecular weight of co-polyamino acid AB17 is    4500 g/mol.-   HPLC-aqueous SEC (calibrant PEG): Mn=3500 g/mol.

EXAMPLE AB18 Co-Polyamino Acid AB18—Sodium poly-L-glutamate Modified atOne of its Ends by Molecule AA7 and Having a Number Average MolecularWeight (Mn) of 3700 g/mol

A sodium poly-L-glutamate modified at one of its ends by molecule AA7 isobtained by polymerization of the γ-methyl N-carboxyanhydride ofglutamic acid (25.0 g, 133.6 mmol) using the hydrochloride salt ofmolecule AA7 (2.80 g, 4.32 mmol) as initiator and by carrying out adeprotection of the methyl esters by using a solution of hydrochloricacid at 37% according to the method described in the patent applicationFR-A-2 801 226.

-   Dry extract: 44.3 mg/g.-   DP (estimated by ¹H NMR)=22, thus i=0.045.-   The calculated average molecular weight of co-polyamino acid AB18 is    3896 g/mol.-   HPLC-aqueous SEC (calibrant PEG): Mn=3700 g/mol.

EXAMPLE AB19 Co-Polyamino Acid AB19—Sodium poly-L-glutamate Modified atOne of its Ends by Molecule AA6 and Having a Number Average MolecularWeight (Mn) of 10,500 g/mol

By a method similar to the one used for the preparation of co-polyaminoacid AB14 applied to the hydrochloride salt of molecule AA6 (1.64 g,2.99 mmol) and to γ-benzyl-L-glutamate N-carboxyanhydride (49.3 g, 187mmol), a sodium poly-L-glutamate modified at one of its ends by moleculeAA6 is obtained.

-   Dry extract: 23.4 mg/g.-   DP (estimated by ¹H NMR)=65, thus i=0.015.-   The calculated average molecular weight of co-polyamino acid AB19 is    10,293 g/mol.-   HPLC-aqueous SEC (calibrant PEG): Mn=10,500 g/mol.

EXAMPLE AB20 Co-Polyamino Acid AB20—Sodium poly-L-glutamate Capped atOne of its Ends by an Acetyl Group and Modified at One of its Ends byMolecule AA6 and Having a Number Average Molecular Weight (Mn) of 10,400g/mol

The hydrochloride salt of molecule AA6 (0.545 g, 1.00 mmol), chloroform(10 mL), molecular mesh 4 Å (3 g) as well as the ion exchange resinAmberlite IRN 150 (3 g) are introduced successively into a suitablecontainer. After 1 h of stirring on rollers, the medium is filtered andthe resin is rinsed with chloroform. The mixture is evaporated, thenco-evaporated with toluene. The residue is solubilized in anhydrous DMF(10 mL) to be used directly in the polymerization reaction.

γ-Benzyl-L-glutamate N-carboxyanhydride (17.0 g, 64.6 mmol) is placedunder vacuum for 30 min in a round-bottom flask dried in the oven, thenanhydrous DMF (30 mL) is introduced. The mixture is stirred under argonuntil the solubilization is complete, cooled at 4° C., then the solutionof molecule AA6 prepared as described above is introduced rapidly. Themixture is stirred between 4° C. and ambient temperature for 2 days,then precipitated in diisopropyl ether (0.6 L). The precipitate isrecovered by filtration, washed two times with diisopropyl ether (40mL), then dried to yield a white solid which is dissolved in 80 mL ofTHF. DIPEA (1.7 mL, 9.8 mmol) then acetic anhydride (0.9 mL, 9.5 mmol)are added successively to this solution. After stirring overnight atambient temperature, the solution is poured slowly into diisopropylether (480 mL) in 30 min and under stirring. After 1 h of stirring, theprecipitate is filtered, washed two times with diisopropyl ether (80mL), then dried under a vacuum at 30° C. to yield apoly(gamma-benzyl-L-glutamic acid) capped at one end by an acetyl groupand modified at the other end by molecule AA6 in the form of a whitesolid.

The solid is diluted in TFA (65 mL), and then a solution of hydrobromicacid (HBr) at 33% in acetic acid (45 mL, 257.0 mmol) is added dropwiseand at 4° C. The solution is stirred for 2 h at ambient temperature,then poured dropwise into a 1:1 (v/v) mixture of diisopropyl ether/waterand under stirring (780 mL). After 2 h of stirring, the heterogeneousmixture is allowed to stand overnight. The white precipitate isrecovered by filtration, washed successively with a 1:1 (v/v) mixture ofdiisopropyl ether and water (70 mL), then with water (70 mL). The solidobtained is solubilized in water (300 mL) by adjusting the pH to 7 byaddition of an aqueous sodium hydroxide solution 10 N, then an aqueoussodium hydroxide solution 1 N. After solubilization, the theoreticalconcentration is adjusted to 20 g/L theoretical by addition of water toobtain a final volume of 440 mL. The mixture is filtered through a 0.45μm filter, then purified by ultrafiltration against a solution of NaCl0.9%, then water until the conductimetry of the permeate is less than 50μS/cm. The solution of co-polyamino acid is then concentrated toapproximately 30 g/L theoretical and the pH is adjusted to 7.0. Theaqueous solution is filtered through a 0.2 μm filter and stored at 4° C.

-   Dry extract: 21.5 mg/g.-   DP (estimated by ¹H NMR)=60, thus i=0.017.-   The calculated average molecular weight of co-polyamino acid AB20 is    9619 g/mol.-   HPLC-aqueous SEC (calibrant PEG): Mn=10,400 g/mol.    BA: Synthesis of the Hydrophobic Molecules in which p=2

The radicals are represented in the following table by the correspondinghydrophobic molecule after grafting onto the co-polyamino acid.

TABLE 1D list of the hydrophobic molecules synthesized according to theinvention in which p = 2. No Structure of the hydrophobic moleculebefore grafting onto the co-polyamino acid BA1

BA2

BA3

BA4

BA5

BA6

BA7

EXAMPLE BA1 Molecule BA1 Molecule B 1: Product Obtained by the ReactionBetween decanoic acid and L-proline

Dicyclohexyl carbodiimide (DCC) (16.29 g, 78.96 mmol) andN-hydroxysuccinimide (NHS) (9.09 g, 78.96 mmol) are added successivelyto a solution of decanoic acid (14.28 g, 82.91 mmol) in THF (520 mL) at0° C. After 60 h of stirring at ambient temperature, the medium iscooled at 0° C. for 20 min, filtered through a sintered filter.L-Proline (10 g, 86.86 mmol), diisopropylethylamine (DIPEA) (68.8 mL)and water (60 mL) are added to the filtrate. After 24 h of stirring atambient temperature, the mixture is diluted with water (300 mL). Theaqueous phase is washed with ethyl acetate (2×250 mL), acidified to pH˜1 with an aqueous HCl solution 1 N, then extracted with dichloromethane(3×150 mL). The combined organic phases are dried over Na₂SO₄, filtered,concentrated under a vacuum, and the residue is purified bychromatography on silica gel (cyclohexane, ethyl acetate).

Yield: 14.6 g (69%).

¹H NMR (CDCl₃, ppm): 0.87 (3H); 1.26 (12H); 1.65 (2H); 2.02 (3H); 2.34(2H); 2.41 (1H); 3.48 (1H); 3.56 (1H); 4.58 (1H).

LC/MS (ESI): 270.2; (calculated ([M+H]⁺): 270.4).

Molecule B2: Product Obtained by the Reaction Between Molecule B1 andL-lysine

By a method similar to the one used for the preparation of molecule Blapplied to molecule B1 (14.57 g, 54.07 mmol) and to L-lysine (4.15 g,28.39 mmol), a yellow oil is obtained.

Yield: 16.4 g (93%).

¹H NMR ¹H (CDCl₃, ppm): 0.88 (6H); 1.26 (24H); 1.35-1.65 (8H); 1.85-2.35(12H); 2.53 (0.2H); 2.90 (0.8H); 3.45-3.75 (5H); 4.50-4.70 (3H); 7.82(1H).

LC/MS (ESI): 649.6; (calculated ([M+H]⁺): 649.9).

Molecule B3: Product Obtained by Reaction Between Molecule B2 andBoc-ethylenediamine

DIPEA (8.80 mL) and 2-(1H-benzotriazol-1-yl)-1,1,3,3-tetramethyluroniumtetrafluoroborate (TBTU, 8.52 g, 26.54 mmol) at ambient temperature areadded to a solution of molecule B2 (16.4 g, 25.27 mmol) in THF (170 mL).After 30 min of stirring, Boc-ethylenediamine (4.45 g, 27.8 mmol) isadded. After stirring at ambient temperature for 2 h, the solvent isevaporated at reduced pressure and the residue is diluted with ethylacetate (400 mL). The organic phase is washed with water (250 mL), asaturated aqueous solution of NaHCO₃ (250 mL), a 1 N aqueous HClsolution (250 mL), a saturated aqueous solution of NaCl (250 mL) anddried over Na₂SO₄. After filtration and concentration under a vacuum,the residue obtained is purified by chromatography on silica gel (ethylacetate, methanol) to yield a colorless oil.

Yield: 12.8 g (64%).

¹H NMR (CDCl₃, ppm): 0.87 (6H); 1.25-1.60 (42H); 1.80-2.05 (4H);2.15-2.45 (9H); 3.10-3.75 (10H); 4.30 (1H); 4.50 (2H); 5.50 (0.6H); 5.89(0.2H); 6.15 (0.2H); 7.03 (1H); 7.47 (1H).

LC/MS (ESI): 791.8; (calculated ([M+H]⁺): 792.1).

Molecule BA1

To a solution of molecule B3 (12.78 g, 16.15 mmol) in dichloromethane(110 mL) at 5° C., an HCl solution 4 N in dioxane (20.2 mL) is added.After 20 h of stirring at 5° C., the medium is concentrated under avacuum. The residue obtained is dissolved in methanol and evaporatedunder a vacuum, this operation being repeated 4 times to yield a whitesolid of molecule BA1 in the form of a hydrochloride salt.

Yield: 11.4 g (97%).

¹H NMR (DMSO-d₆, ppm): 0.85 (6H); 1.25-1.50 (33H); 1.57 (1H); 1.70-2.40(12H); 2.82 (2H); 3.00 (2H); 3.25-3.70 (6H); 4.05-4.50 (3H); 7.75-8.45(6H).

LC/MS (ESI): 691.6; (calculated ([M+H]⁺): 692.0).

EXAMPLE BA2 Molecule BA2 Molecule B4: Product Obtained by the ReactionBetween Lauric Acid and L-proline

By a method similar to the one used for the preparation of molecule B1,applied to lauric acid (31.83 g, 157.9 mmol) and to L-proline (20 g,173.7 mmol), a yellow oil is obtained.

Yield: 34.3 g (73%).

¹H NMR (CDCl3, ppm): 0.87 (3H); 1.26 (16H); 1.70 (2H); 1.90-2.10 (3H);2.35 (2H); 2.49 (1H); 3.48 (1H); 3.56 (1H); 4.60 (1H).

LC/MS (ESI): 298.2; (calculated ([M+H]⁺): 298.4).

Molecule B5: Product Obtained by the Reaction Between Molecule B4 andL-lysine

By a method similar to the one used for the preparation of molecule Blapplied to molecule B4 (33.72 g, 113.36 mmol) and to L-lysine (8.70 g,59.51 mmol), a white solid is obtained.

Yield: 26.2 g (66%).

¹H NMR (CDCl₃, ppm): 0.88 (6H); 1.26 (32H); 1.35-1.65 (8H); 1.85-2.35(15H); 2.87 (1H); 3.40-3.75 (5H); 4.50-4.75 (3H); 7.87 (1H).

LC/MS (ESI): 705.6; (calculated ([M+H]⁺): 706.0).

Molecule B6: Product Obtained by Reaction Between Boc-ethylenediamineand Molecule B5

By a method similar to the one used for the preparation of moleculeB3applied to molecule B5 (25.74 g, 36.51 mmol) and toBoc-ethylenediamine (6.43 g, 40.16 mmol), a colorless oil is obtained.

Yield: 30.9 g (quantitative).

¹H NMR (CDCl₃, ppm): 0.88 (6H); 1.35-1.65 (50H); 1.85-2.35 (13H);3.05-3.75 (10H); 4.25-4.65 (3H); 5.50 (0.4H); 5.88 (0.2H); 6.16 (0.2H);7.08 (1H); 7.26 (1H); 7.49 (0.2H).

LC/MS (ESI): 847.8; (calculated ([M+H]⁺): 848.2).

Molecule BA2

After a method similar to the one used for the preparation of moleculeBA1 applied to molecule B6 (30.9 g, 36.47 mmol), the residue obtainedafter concentration under a vacuum is dissolved in methanol andevaporated under a vacuum, this operation being repeated 4 times toyield a white solid of molecule BA2 in the form of a hydrochloride saltafter drying at reduced pressure.

Yield: 27.65 g (97%).

¹H NMR (DMSO-d₆, ppm): 0.85 (6H); 1.10-2.40 (54H); 2.75-3.15 (4H);3.25-3.60 (6H); 4.05-4.50 (3H); 7.50-8.50 (6H).

LC/MS (ESI): 747.6; (calculated ([M+H]⁺): 748.1).

EXAMPLE BA3 Molecule BA3 Molecule B7: Product Obtained by the ReactionBetween Myristic Acid and L-proline

By a method similar to the one used for the preparation of molecule B1,applied to myristic acid (18.93 g, 82.91 mmol) and to L-proline (10 g,86.86 mmol), a yellowish oil is obtained.

Yield: 20 g (78%).

¹H NMR (CDCl₃, ppm): 0.88 (3H); 1.28 (20H); 1.70 (2H); 1.90-2.10 (3H);2.36 (2H); 2.51 (1H); 3.47 (1H); 3.56 (1H); 4.61 (1H).

LC/MS (ESI): 326.2; (calculated ([M+H]⁺): 326.6).

Molecule B8: Product Obtained by the Reaction Between Molecule B7 andL-lysine

By a method similar to the one used for the preparation of molecule Blapplied to molecule B7 (20.02 g, 61.5 mmol) and to L-lysine (4.72 g,32.29 mmol), a white solid is obtained.

Yield: 12.3 g (53%).

¹H NMR (DMSO-d₆, ppm): 0.85 (6H); 1.26 (40H); 1.35-1.50 (6H); 1.50-2.10(10H); 2.10-2.25 (4H); 3.01 (2H); 3.31-3.55 (4H); 4.10-4.40 (3H); 7.68(0.6H); 7.97 (1H); 8.27 (0.4H); 12.50 (1H).

LC/MS (ESI): 761.8; (calculated ([M+H]⁺): 762.1).

Molecule B9: Product Obtained by the Reaction BetweenBoc-ethylenediamine and Molecule B8

By a method similar to the one used for the preparation of moleculeB3applied to molecule B8 (12 g, 15.77 mmol) and to Boc-ethylenediamine(3.03 g, 18.92 mmol), a colorless oil is obtained after purification bychromatography column on silica gel (ethyl acetate, methanol).

Yield: 12.5 g (88%).

¹H NMR (DMSO-d₆, ppm): 0.85 (6H); 1.20-1.55 (55H); 1.50-2.25 (14H);2.95-3.10 (6H); 3.31-3.55 (4H); 4.10-4.40 (3H); 6.74 (1H); 7.60-8.25(3H).

LC/MS (ESI): 904.1; (calculated ([M+H]⁺): 904.3).

Molecule BA3

After a method similar to the one used for the preparation of moleculeBA1 applied to molecule B9 (12.5 g, 13.84 mmol), the residue obtainedafter concentration under a vacuum is dissolved in methanol andevaporated under a vacuum, this operation being repeated 4 times toyield a white solid of molecule BA3 in the form of a hydrochloride saltafter drying at reduced pressure.

Yield: 9.2 g (79%).

¹H NMR (DMSO-d₆, ppm): 0.85 (6H); 1.10-1.65 (48H); 1.70-2.35 (12H); 2.85(2H); 3.01 (2H); 3.25-3.65 (6H); 4.10-4.50 (3H); 7.70-8.40 (6H).

LC/MS (ESI): 803.9; (calculated ([M+H]⁺): 804.2).

EXAMPLE BA4 Molecule BA4 Molecule B10: Product Obtained by the ReactionBetween Molecule B8 and Boc-1-amino-4,7,10-trioxa-13-tridecane amine

By a method similar to the one used for the preparation of molecule B3applied to molecule B8 (29.80 g, 39.15 mmol) and toBoc-1-amino-4,7,10-trioxa-13-tridecane amine (15.05 g, 46.96 mmol), athick colorless oil is obtained.

Yield: 25.3 g (61%).

¹H NMR (DMSO-d₆, ppm): 0.85 (6H); 1.25-2.35 (75H); 2.85-3.20 (6H);3.25-3.65 (16H); 4.10-4.45 (3H); 6.38 (0.1H); 6.72 (0.9H); 7.50-8.25(3H).

LC/MS (ESI): 1064.2; (calculated ([M+H]⁺): 1064.5).

Molecule BA4

After a method similar to the one used for the preparation of moleculeBA1 applied to molecule B10 (25.3 g, 23.8 mmol), the residue obtainedafter concentration under a vacuum is dissolved in methanol andevaporated under a vacuum, this operation being repeated 4 times toyield a white solid of molecule BA4 in the form of a hydrochloride saltafter drying at reduced pressure.

Yield: 20.02 g (84%).

¹H NMR (DMSO-d₆, ppm): 0.85 (6H); 1.15-2.35 (66H); 2.80-3.20 (6H);3.30-3.65 (16H); 4.10-4.45 (3H); 7.55-8.60 (6H).

LC/MS (ESI): 964.9; (calculated ([M+H]⁺): 964.6).

EXAMPLE BA5 Molecule BA5 Molecule B11: Product Obtained by ReactionBetween Molecule A1 and L-lysine

By a method similar to the one used for the preparation of molecule B1applied to molecule A1 (19.10 g, 54.02 mmol) and to L-lysine (4.15 g,28.36 mmol), an oily residue is obtained after concentration of thereaction medium at reduced pressure. This residue is diluted in water(150 mL), washed with ethyl acetate (2×75 mL), then the aqueous phase isacidified until the pH is 1 by slow addition of HCl 6 N. The product isextracted 3 times with dichloromethane, the organic phase is dried overNa₂SO₄, then filtered and concentrated at reduced pressure to yield 11.2g of yellow oily residue. In parallel, the previous preceding organicphase of ethyl acetate is washed with an aqueous HCl solution 2 N (2×75mL), a saturated aqueous solution of NaCl (75 mL), dried over Na₂SO₄,filtered and concentrated to yield 10.2 g of yellow oily residue. Awhite solid is obtained after recrystallization of each one of theseresidues in acetone.

Yield: 11.83 g (54%).

¹H NMR (CDCl₃, ppm): 0.87 (6H); 1.06-2.44 (70H); 2.78-2.96 (1H);3.35-3.75 (5H); 4.28-4.43 (0.1H); 4.43-4.52 (0.2H); 4.52-4.61 (1.8H);4.61-4.75 (0.9H); 7.74-8.02 (2H).

LC/MS (ESI): 818.0; (calculated ([M+H]⁺): 818.7).

Molecule B12: Product Obtained by Coupling Between Molecule B11 andBoc-ethylenediamine

By a method similar to the one used for the preparation of moleculeB3applied to molecule B11 (18.00 g, 22.02 mmol) in solution in THF andto Boc-ethylenediamine (4.23 g, 26.43 mmol), a white solid is obtainedafter recrystallization two times in acetonitrile.

Yield: 17.5 g (83%).

¹H NMR (DMSO-d₆, ppm): 0.85 (6H); 1.15-2.29 (79H); 2.92-3.12 (6H);3.30-3.59 (4H); 4.06-4.13 (0.65H); 4.16-4.29 (2H); 4.38-4.42 (0.35H);6.71-6.76 (1H); 7.60-7.69 (1.3H); 7.76-7.81 (0.65H); 7.93-7.97 (0.35H);8.00-8.04 (0.35H); 8.10-8.17 (0.35H).

LC/MS (ESI): 960.4; (calculated ([M+H]⁺): 960.8).

Molecule BA5

By a method similar to the one used for the preparation of molecule BA1applied to molecule B12 (24.4 g, 25.43 mmol), the residue obtained afterconcentration under a vacuum is solubilized in dichloromethane (150 mL),the organic phase is washed 2 times with an aqueous sodium hydroxidesolution 2 N (90 mL). Acetonitrile (120 mL) is added, and thedichloromethane is eliminated by concentration at reduced pressure. Themedium is then left to stand for 72 h and a white solid is obtainedafter filtration and rinsing with acetonitrile, followed by drying atreduced pressure. This operation is repeated 4 times.

Yield: 14.28 g (65%).

¹H NMR (DMSO-d₆, ppm): 0.85 (6H); 1.06-2.32 (70H); 2.53-2.63 (2H);2.89-3.61 (10H); 4.04-4.43 (3H); 7.55-7.62 (0.65H); 7.65-7.72 (0.65H);7.80 (0.65H); 7.91 (0.35H); 8.03 (0.35H); 8.14-8.23 (0.35H).

LC/MS (ESI): 860.0; (calculated ([M+H]⁺): 860.8).

EXAMPLE BA6 Molecule BA6 Molecule B13: Product Obtained by the ReactionBetween N-(tert-butoxycarbonyl)-1,6-diaminohexane and Molecule B8

By a method similar to the one used for the preparation of moleculeB3applied to molecule B8 (10 g, 13.14 mmol) and toN-(tert-butoxycarbonyl)-1,6-diaminohexane (3.41 g, 15.77 mmol) indichloromethane, a white solid is obtained after recrystallization inacetonitrile.

Yield: 10.7 g (85%).

¹H NMR (CDCl₃, ppm): 0.88 (6H); 1.17-2.40 (79H); 3.00-3.71 (10H);4.26-4.58 (3H); 4.67 (1H); 6.74 (1H); 7.34-7.49 (2H).

LC/MS (ESI): 959.9; (calculated ([M+H]⁺): 959.8).

Molecule BA6

After a method similar to the one used for the preparation of moleculeBA1 applied to molecule B13 (10.5 g, 10.94 mmol), an aqueous NaOHsolution 2 N is added dropwise to the reaction medium cooled to 0° C.The aqueous phase is extracted with dichloromethane, then the organicphase is washed 3 times with an aqueous solution of NaCl 5%. Afterdrying over Na₂SO₄, the organic phase is filtered, concentrated under avacuum, and the residue is recrystallized in acetonitrile.

Yield: 5.4 g (58%).

¹H NMR (CDCl₃, ppm): 0.88 (6H); 1.19-2.40 (72H); 2.67 (2H); 3.03-3.70(8H); 4.26-4.57 (3H); 6.71 (1H); 7.39-7.49 (2H).

LC/MS (ESI): 859.8; (calculated ([M+H]⁺): 859.7).

EXAMPLE BA7 Molecule BA7 Molecule B14: Product Obtained by CouplingBetween Molecule B7 and 2,3-diaminopropionic acid

By a method similar to the one used for the preparation of molecule B1applied to molecule B7 (80.00 g, 245.78 mmol) and to the dihydrochlorideof 2,3-diaminopropionic acid (22.84 g, 129.04 mmol), a white solid isobtained after recrystallization in acetonitrile.

Yield: 69 g (78%).

¹H NMR (DMSO-d₆, ppm): 0.86 (6H); 1.08-1.38 (40H); 1.40-1.55 (4H);1.68-2.30 (12H); 3.16-3.66 (6H); 4.20-4.39 (3H); 7.67-8.31 (2H); 12.70(1H).

LC/MS (ESI): 719.4; 741.5; (calculated ([M+H]⁺): 719.6; ([M+Na]⁺):741.6).

Molecule B15: Product Obtained by Coupling Between Molecule B14 andBoc-ethylenediamine

By a method similar to the one used for the preparation of molecule B3applied to molecule B14 (32.00 g, 44.50 mmol) in solution indichloromethane and to Boc-ethylenediamine (8.56 g, 53.40 mmol), acolorless oil is obtained after purification by chromatography on silicagel (ethyl acetate, methanol).

Yield: 24.5 g (64%).

¹H NMR (DMSO-d₆, ppm): 0.85 (6H); 1.16-2.42 (65H); 2.89-3.14 (4H);3.17-3.66 (6H); 4.11-4.43 (3H); 6.77 (1H); 7.38-8.23 (3H).

LC/MS (ESI): 861.7; (calculated ([M+H]⁺): 861.7).

Molecule BA7

After a method similar to the one used for the preparation of moleculeBA1 applied to molecule B15 (24.50 g, 28.45 mmol), the reaction mediumis concentrated at reduced pressure, the residue is solubilized indichloromethane, the organic phase is washed with an aqueous NaOHsolution 2 N, dried over Na₂SO₄, filtered and concentrated at reducedpressure. A white solid is obtained after recrystallization inacetonitrile.

Yield: 19.7 g (91%).

¹H NMR (DMSO-d₆, ppm): 0.85 (6H); 1.10-2.40 (58H); 2.51-2.62 (2H);2.90-3.16 (2H); 3.16-3.67 (6H); 4.04-4.47 (3H); 7.33-8.27 (3H).

LC/MS (ESI): 761.5; (calculated ([M+H]⁺): 761.6).

BB: Synthesis of the Co-Polyamino Acids Modified by HydrophobicMolecules in which p=2

Co-Polyamino Acids of Formula VII or VIIa

TABLE 1e list of the co-polyamino acids of formula VII or VIIa accordingto the invention. no co-polyamino acids bearing carboxylate charges andhydrophobic radicals BB1

BB2

BB3

BB4

BB5

BB6

BB7

BB8

BB9

BB10

BB11

BB12

BB13

Co-Polyamino Acids of Formula VII or VIIb

TABLE 1F list of the co-polyamino acids of formula VII or VIIbsynthesized according to the invention. no co-polyamino acids bearingcarboxylate charges and hydrophobic radicals BB14

BB15

BB16

BB17

BB18

BB19

BB20

BB21

BB22

BB23

BB24

BB25

EXAMPLE BB1 Co-Polyamino Acid BB1—Sodium poly-L-glutamate Modified byMolecule BA2 and Having a Number Average Molecular Weight (Mn) of 2400g/mol

Co-polyamino acid BB1-1: poly-L-glutamic acid having a relative numberaverage molecular weight (Mn) 3860 g/mol originating from thepolymerization of γ-benzyl-L-glutamate N-carboxyanhydride initiated byhexylamine.

In a previously oven-heated round-bottom flask, γ-benzyl-L-glutamateN-carboxyanhydride (90.0 g, 342 mmol) is placed under vacuum for 30 min,then anhydrous DMF (465 mL) is introduced. The mixture is then stirredunder argon until the dissolution is complete, cooled to 4° C., thenhexylamine (1.8 mL, 14 mmol) is introduced rapidly. The mixture isstirred between 4° C. and ambient temperature for 2 days. The reactionmixture is then heated at 65° C. for 4 h, cooled to ambient temperature,then poured dropwise into cold diisopropyl ether (6 L) under stirring.The white precipitate is recovered by filtration, washed withdiisopropyl ether (500 mL, then 250 mL), then dried under a vacuum at30° C. to yield poly(γ-benzyl-L-glutamic acid) (PBLG).

A hydrobromic acid solution (HBr) at 33% in acetic acid (135 mL, 0.77mol) is added dropwise to a solution of PBLG (42.1 g) in trifluoroaceticacid (TFA, 325 mL). The mixture is stirred at ambient temperature for 2h, then poured dropwise into a 1:1 (v/v) mixture of diisopropyl etherand water under stirring (1.6 L). After 1 h 30 of stirring, theheterogeneous mixture is allowed to stand overnight. The whiteprecipitate is recovered by filtration, washed with a 1:1 (v/v) mixtureof diisopropyl ether and water (200 mL).

The solution obtained is then solubilized in water (1 L) by adjustingthe pH to 7 by addition of an aqueous sodium hydroxide solution 10 N,then an aqueous sodium hydroxide solution 1 N. After solubilization, thetheoretical concentration is adjusted to 25 g/L theoretical by additionof water to obtain a final volume of 1.5 L.

The solution is filtered through a 0.45 μm filter, then purified byultrafiltration against a solution of NaCl 0.9%, then water until theconductimetry of the permeate is less than 50 μS/cm.

The aqueous solution is then acidified by addition of a solution ofhydrochloric acid 37% until a pH of 2 is reached. After 4 h of stirring,the precipitate obtained is filtered, then dried under a vacuum at 30°C. to yield a poly-L-glutamic acid having a number average molecularweight (Mn) 3860 g/mol with respect to a polyoxyethylene standard (PEG).

Co-Polyamino Acid BB1

The co-polyamino acid BB1-1 (10.0 g) is solubilized in DMF (700 mL) at30-40° C., then cooled to 0° C. The hydrochloride salt of molecule BA2(2.95 g, 3.8 mmol) is suspended in DMF (45 mL) and triethylamine (0.39g, 3.8 mmol) is then added to this suspension, then the mixture isheated slightly under stirring until the dissolution is complete.N-Methylmorpholine (NMM, 7.6 g, 75 mmol) in DMF (14 mL) and ethylchloroformate (ECF, 8.1 g, 75 mmol) are added to the solution ofco-polyamino acid at 0° C. After 10 min at 0° C., the solution ofmolecule BA2 is added, and the mixture is maintained at 30° C. for 1 h.The reaction mixture is poured dropwise into 6 L of water containingNaCl at 15 wt % and HCl (pH 2), then allowed to stand overnight. Theprecipitate is collected by filtration, washed with the solution ofsodium chloride at pH 2 (1 L) and dried under a vacuum for approximately1 h. The white solid obtained is taken up in water (600 mL), and the pHis adjusted to 7 by slow addition of an aqueous NaOH solution 1 N. Thevolume is adjusted to 700 mL by addition of water. After filtrationthrough a 0.45 μm filter, the clear solution obtained is purified byultrafiltration against a solution of NaCl 0.9%, then water until theconductimetry of the permeate is less than 50 μS/cm. After removal, thesolution is filtered through a 0.2 μm filter and stored at 2-8° C.

-   Dry extract: 19.7 mg/g.-   DP (estimated based on ¹H NMR): 23.-   Based on ¹H NMR: i=0.05.-   The calculated average molecular weight of co-polyamino acid BB1 is    4350 g/mol.-   HPLC-aqueous SEC (calibrant PEG): Mn=2400 g/mol.

EXAMPLE BB2 Co-Polyamino Acid BB2—Sodium poly-L-glutamate Modified byMolecule BA2 and Having a Number Average Molecular Weight (Mn) of 4900g/mol

A poly-L-glutamic acid having a number average molecular (Mn) 4100 g/mol(5.0 g) obtained by a method similar to the one used for the preparationof co-polyamino acid BB1-1 is solubilized in DMF (205 mL) at 30-40° C.,then maintained at this temperature. In parallel, the hydrochloride saltof molecule BA2 (1.44 g, 1.84 mmol) is suspended in DMF (10 mL), andtriethylamine (0.19 g, 1.84 mmol) is added, then the mixture is heatedslightly under stirring until the dissolution is complete. To thesolution of co-polyamino acid in DMF, NMM (3.7 g, 36.7 mmol), thesolution of molecule BA2, then the N-oxide of 2-hydroxypyridine (HOPO,0.31 g, 2.76 mmol) are added successively. The reaction medium is thencooled to 0° C., then EDC (0.53 g, 2.76 mmol) is added, and the mixtureis brought to ambient temperature in 3 h. The reaction medium is poureddropwise into 1.55 L of water containing NaCl at 15 wt % and HCl (pH 2)under stirring. At the end of the addition, the pH is readjusted to 2with an HCl solution 1 N, and the suspension is allowed to standovernight. The precipitate is collected by filtration, then rinsed with100 mL of water. The white solid obtained is solubilized in 200 mL ofwater by slow addition of an aqueous NaOH solution 1 N until the pH is 7under stirring, then the solution is filtered through a 0.45 μm filter.The clear solution obtained is purified by ultrafiltration against asolution of NaCl 0.9%, then water until the conductimetry of thepermeate is less than 50 μS/cm. The solution obtained is filteredthrough a 0.2 μm filter and stored at 2-8° C.

-   Dry extract: 16.3 mg/g.-   DP (estimated based on ¹H NMR): 21.-   Based on ¹H NMR: i=0.047.-   The calculated average molecular weight of co-polyamino acid BB2 is    3932 g/mol.-   HPLC-aqueous SEC (calibrant PEG): Mn=4900 g/mol.

EXAMPLE BB3 Co-Polyamino Acid BB3—Sodium poly-L-glutamate Modified byMolecule BA2 and Having a Number Average Molecular Weight (Mn) of 6400g/mol

Co-polyamino acid BB3-1: poly-L-glutamic acid having a number averagemolecular weight (Mn) 17,500 g/mol originating from the polymerizationof γ-methyl-L-glutamate N-carboxyanhydride initiated by L-leucinamide.

A poly-L-glutamic acid having a number average molecular weight (Mn)17,500 g/mol with respect to a methyl polymethacrylate (PMMA) standardis obtained by polymerization of the γ-methyl N-carboxyanhydride ofglutamic acid using L-leucinamide as initiator and by carrying out adeprotection of the methyl esters by using a solution of hydrochloricacid at 37% according to the method described in the patent applicationFR-A-2 801 226.

By a method similar to the one used for the preparation of co-polyaminoacid BB2 applied to the hydrochloride salt of molecule BA2 (3.23 g, 4.1mmol) and to co-polyamino acid BB3-1 (11 g), a sodium poly-L-glutamatemodified by molecule BA2 is obtained.

-   Dry extract: 27.5 mg/g.-   DP (estimated based on ¹H NMR): 34.-   Based on ¹H NMR: i=0.049.-   The calculated average molecular weight of co-polyamino acid BB3 is    6405 g/mol.-   HPLC-aqueous SEC (calibrant PEG): Mn=6400 g/mol.

EXAMPLE BB4 Co-Polyamino Acid BB4—Sodium poly-L-glutamate Modified byMolecule BA2 and Having a Number Average Molecular Weight (Mn) of 10,500g/mol

By a method similar to the one used for the preparation of co-polyaminoacid BB2 applied to the hydrochloride salt of molecule BA2 (5 g, 6.35mmol) and to a poly-L-glutamic acid having a number average molecularweight Mn=10,800 g/mol (21.7 g) obtained by a method similar to the oneused for the preparation of copolyamino acid BB1-1, a sodiumpoly-L-glutamate modified by molecule BA2 is obtained.

-   Dry extract: 28.2 mg/g.-   DP (estimated based on ¹H NMR): 65.-   Based on ¹H NMR: i=0.04.-   The calculated average molecular weight of co-polyamino acid BB4 is    11,721 g/mol.-   HPLC-aqueous SEC (calibrant PEG): Mn=10,500 g/mol.

EXAMPLE BB5 Co-Polyamino Acid BB5—Sodium poly-L-glutamate Capped at Oneof its Ends by an Acetyl Group and Modified by Molecule BA2 and Having aNumber Average Molecular Weight (Mn) of 3600 g/mol

Co-polyamino acid BB5-1: poly-L-glutamic acid of Mn 3700 g/moloriginating from the polymerization of γ-benzyl-L-glutamateN-carboxyanhydride initiated by hexylamine and capped at one of its endsby an acetyl group.

In a round-bottom flask dried in the oven, γ-benzyl-L-glutamateN-carboxyanhydride (100.0 g, 380 mmol) is placed under a vacuum for 30min, then anhydrous DMF (250 mL) is introduced. The mixture is thenstirred under argon until the dissolution is complete, cooled to 4° C.,then hexylamine (2.3 mL, 17 mmol) is introduced rapidly. The mixture isstirred between 4° C. and ambient temperature for 2 days, thenprecipitated in diisopropylene (3.4 L). The precipitate is recovered byfiltration, washed 2 times with diisopropyl ether (225 mL), then driedto yield a white solid which is dissolved in 450 mL of THF. To thissolution, N,N-diisopropylethylamine (DIPEA, 31 mL, 176 mmol), thenacetic anhydride (17 mL, 176 mmol) are added successively. Afterstirring overnight at ambient temperature, the solution is poured slowlyinto diisopropyl ether (3 L) for a duration of 30 min and understirring. After 1 h of stirring, the precipitate is filtered, washed twotimes with diisopropyl ether (200 mL), then dried under a vacuum at 30°C. to yield a poly(γ-benzyl-L-glutamic acid) capped at one of its endsby an acetyl group.

A solution of hydrobromic acid (HBr) at 33% in acetic acid (235 mL, 1.34mol) is added dropwise to a solution of the capped co-polyamino acid (72g) in trifluoroacetic acid (TFA, 335 mL) at 4° C. The mixture is stirredat ambient temperature for 3 h 30, then poured dropwise onto a 1:1 (v/v)mixture of diisopropyl ether and water under stirring (4 L). After 2 hof stirring, the heterogeneous mixture is allowed to stand overnight.The white precipitate is recovered by filtration, washed with a 1:1(v/v) mixture of diisopropyl ether and water (340 mL), then with water(340 mL). The solid obtained is then solubilized in water (1.5 L) byadjusting the pH to 7 by addition of an aqueous sodium hydroxidesolution 10 N, then an aqueous sodium hydroxide solution 1 N. Aftersolubilization, the theoretical concentration is adjusted to 20 g/Ltheoretical by addition of water to obtain a final volume of 2.1 L. Thesolution is filtered through a 0.45 μm filter, then purified byultrafiltration against a solution of NaCl 0.9%, then water until theconductimetry of the permeate is less than 50 μS/cm. The solution ofco-polyamino acid is then concentrated until a final volume of 1.8 L isobtained. The aqueous solution is then acidified by addition of solutionof hydrochloric acid 37% until a pH of 2 is obtained. After 4 h ofstirring, the precipitate obtained is filtered, washed with water (330mL), then dried under a vacuum at 30° C. to yield a poly-L-glutamic acidhaving a number average molecular weight (Mn) 3700 g/mol with respect toa polyoxyethylene standard (PEG).

Co-Polyamino Acid BB5

By a method similar to the one used for the preparation of co-polyaminoacid BB2 applied to the hydrochloride salt of molecule BA2 (6.92 g, 8.8mmol) and to co-polyamino acid BB5-1 (30.0 g), a sodium poly-L-glutamatecapped at one of its ends by an acetyl group and modified by moleculeBA2 is obtained.

-   Dry extract: 29.4 mg/g.-   DP (estimated based on ¹H NMR): 23.-   Based on ¹H NMR: i=0.042.-   The calculated average molecular weight of co-polyamino acid BB5 is    4302 g/mol.-   HPLC-aqueous SEC (calibrant PEG): Mn=3600 g/mol.

EXAMPLE BB6 Co-Polyamino Acid BB6—Sodium poly-L-glutamate Capped at Oneof its Ends by an Acetyl Group and Modified by Molecule BA2 and Having aNumber Average Molecular Weight (Mn) of 4100 g/mol

By a method similar to the one used for the preparation of co-polyaminoacid BB2 applied to the hydrochloride salt of molecule BA2 (5.8 g, 7.4mmol) and to a poly-L-glutamic acid having a number average molecularweight Mn=3800 g/mol (25 g) obtained by a method similar to the one usedfor the preparation of co-polyamino acid BB5-1 using ammonia instead ofhexylamine, a sodium poly-L-glutamate capped at one of its ends by anacetyl group and modified by molecule BA2 is obtained.

-   Dry extract: 27.6 mg/g.-   DP (estimated based on ¹H NMR): 24.-   Based on ¹H NMR: i=0.04.-   The calculated average molecular weight of co-polyamino acid BB6 is    4387 g/mol.-   HPLC-aqueous SEC (calibrant PEG): Mn=4100 g/mol.

EXAMPLE BB7 Co-Polyamino Acid BB7—Sodium poly-L-glutamate Modified byMolecule BA2 and Having a Number Average Molecular Weight (Mn) of 4200g/mol

By a method similar to the one used for the preparation of co-polyaminoacid BB2 applied to the hydrochloride acid of molecule BA2 (7.07 g, 9.0mmol) and to a poly-L-glutamic acid having a number average molecularweight Mn=3600 g/mol (30.0 g) obtained by a method similar to the oneused for the preparation of co-polyamino acid BB1-1, a sodiumpoly-L-glutamate modified by molecule BA2 is obtained.

-   Dry extract: 28.3 mg/g.-   DP (estimated based on ¹H NMR): 22.-   Based on ¹H NMR: i=0.042.-   The calculated average molecular weight of co-polyamino acid BB7 is    4039 g/mol.-   HPLC-aqueous SEC (calibrant PEG): Mn=4200 g/mol.

EXAMPLE BB8 Co-Polyamino Acid BB8—Sodium poly-L-glutamate Modified byMolecule BA2 and Having a Number Average Molecular Weight (Mn) of 5200g/mol

By a method similar to the one used for the preparation of co-polyaminoacid BB2 applied to the hydrochloride salt of molecule BA2 (0.85 g, 1.1mmol) and to a poly-L-glutamic acid having a number average molecularweight Mn=4100 g/mol (5.0 g) obtained by a method similar to the oneused for the preparation of co-polyamino acid BB1-1, a sodiumpoly-L-glutamate modified by molecule BA2 is obtained.

-   Dry extract: 28.6 mg/g.-   DP (estimated based on ¹H NMR): 21.-   Based on ¹H NMR: i=0.026.-   The calculated average molecular weight of co-polyamino acid BB8 is    3620 g/mol.-   HPLC-aqueous SEC (calibrant PEG): Mn=5200 g/mol.

EXAMPLE BB9 Co-Polyamino Acid BB9—Sodium poly-L-glutamate Modified byMolecule BA3 and Having a Number Average Molecular Weight (Mn) of 4700g/mol

By a method similar to the one used for the preparation of co-polyaminoacid BB2 applied to the hydrochloride salt of molecule BA3 (3.05 g, 3.6mmol) and to a poly-L-glutamic acid having a number average molecularweight Mn=4100 g/mol (10.0 g) obtained by a method similar to the oneused for the preparation of co-polyamino acid BB1-1, a sodiumpoly-L-glutamate modified by molecule BA3 is obtained.

-   Dry extract: 28.6 mg/g.-   DP (estimated based on ¹H NMR): 26.-   Based on ¹H NMR: i=0.05.-   The calculated average molecular weight of co-polyamino acid BB9 is    4982 g/mol.-   HPLC-aqueous SEC (calibrant PEG): Mn=4700 g/mol.

EXAMPLE BB10 Co-Polyamino Acid BB10—Sodium poly-L-glutamate Modified byMolecule BA3 and Having a Number Average Molecular Weight (Mn) of 4200g/mol

By a method similar to the one used for the preparation of co-polyaminoacid BB2 applied to the hydrochloride salt of molecule BA3 (1.90 g, 2.3mmol) and a poly-L-glutamic acid having a number average molecularweight Mn=3500 g/mol (10.0 g) by a method similar to the one used forthe preparation of co-polyamino acid BB1-1, a sodium poly-L-glutamatemodified by molecule BA3 is obtained.

-   Dry extract: 25.9 mg/g.-   DP (estimated based on ¹H NMR): 22.-   Based on ¹H NMR: i=0.029.-   The calculated average molecular weight of co-polyamino acid BB10 is    3872 g/mol.-   HPLC-aqueous SEC (calibrant PEG): Mn=4200 g/mol.

EXAMPLE BB11 Co-Polyamino Acid BB11—Sodium poly-L-glutamate Capped atOne of its Ends by an Acetyl Group and Modified by Molecule BA4 andHaving a Number Average Molecular Weight (Mn) of 3900 g/mol

By a method similar to the one used for the preparation of co-polyaminoacid BB2 applied to the hydrochloride salt of molecule BA4 (2.21 g, 2.2mmol) and to a poly-L-glutamic acid having a number average molecularweight Mn=3700 g/mol (10 g) obtained by a method similar to the one usedfor the preparation of co-polyamino acid BB5-1, a sodiumpoly-L-glutamate capped at one of its ends by an acetyl group andmodified by molecule BA4 is obtained.

-   Dry extract: 28.1 mg/g.-   DP (estimated based on ¹H NMR): 22.-   Based on ¹H NMR: i=0.032.-   The calculated average molecular weight of co-polyamino acid BB11 is    4118 g/mol.-   HPLC-aqueous SEC (calibrant PEG): Mn=3900 g/mol.

EXAMPLE BB12 Co-Polyamino Acid BB12—Sodium poly-L-glutamate Capped atOne of its Ends by an Acetyl Group and Modified by Molecule BA3 andHaving a Number Average Molecular Weight (Mn) of 3900 g/mol

By a method similar to the one used for the preparation of co-polyaminoacid BB2 applied to the hydrochloride salt of molecule BA3 (1.9 g, 2.3mmol) and a poly-L-glutamic acid having a number average molecularweight Mn=3600 g/mol (10 g) obtained by a method similar to the one usedfor the preparation of co-polyamino acid BB5-1, a sodiumpoly-L-glutamate capped at one of its ends by an acetyl group andmodified by molecule BA3 is obtained.

-   Dry extract: 26.7 mg/g.-   DP (estimated based on ¹H NMR): 23.-   Based on ¹H NMR: i=0.03.-   The calculated average molecular weight of co-polyamino acid BB12 is    4145 g/mol.-   HPLC-aqueous SEC (calibrant PEG): Mn=3900 g/mol.

EXAMPLE BB13 Co-Polyamino Acid BB13—Sodium poly-L-glutamate Modified byMolecule BA1 and Having a Number Average Molecular Weight (Mn) of 2800g/mol

By a method similar to the one used for the preparation of co-polyaminoacid BB1 applied to the hydrochloride salt of molecule BA1 (3.65 g, 5mmol) and to a poly-L-glutamic acid having a number average molecularweight Mn=3600 g/mol (10 g) also by a method similar to the one used forthe preparation of co-polyamino acid BB1-1, a sodium poly-L-glutamatemodified by molecule BA1 is obtained.

-   Dry extract: 25.6 mg/g.-   DP (estimated based on ¹H NMR): 25.-   Based on ¹H NMR: i=0.08.-   The calculated average molecular weight of co-polyamino acid BB13 is    5253 g/mol.-   HPLC-aqueous SEC (calibrant PEG): Mn=2800 g/mol.

EXAMPLE BB14 Co-Polyamino Acid BB14—Sodium poly-L-glutamate Modified atOne of its Ends by Molecule BA2 and Having a Number Average MolecularWeight (Mn) of 4020 g/mol

The hydrochloride salt of molecule BA2 (2.12 g, 2.70 mmol), chloroform(40 mL), molecular mesh 4 Å (1.5 g) as well as the ion exchange resinAmberlite IRN 150 (1.5 g) are introduced successively into a suitablecontainer. After 1 h of stirring on rollers, the medium is filtered andthe resin is rinsed with chloroform. The mixture is evaporated, thenco-evaporated with toluene. The residue is solubilized in anhydrous DMF(20 mL) to be used directly in the polymerization reaction.

In a round-bottom flask dried in the oven, γ-benzyl-L-glutamateN-carboxyanhydride (18 g, 68.42 mmol) is placed under a vacuum for 30min, then anhydrous DMF (100 mL) is introduced. The mixture is stirredunder argon until the solubilization is complete, cooled to 4° C., thenthe solution of molecule BA2 prepared as described above is introducedrapidly. The mixture is stirred between 4° C. and ambient temperaturefor 2 days, then heated at 65° C. for 2 h. The reaction mixture is thencooled to ambient temperature, then poured dropwise into diisopropylether (1.2 L) under stirring. The white precipitate is recovered byfiltration, washed two times with diisopropyl ether (100 mL), then driedunder a vacuum at 30° C. to obtain a white solid. The solid is dilutedin TFA (105 mL), and a solution of hydrobromic acid (HBr) at 33% inacetic acid (38 mL, 220 mmol) is then added dropwise and at 0° C. Thesolution is stirred for 2 h at ambient temperature, then poured dropwiseinto a 1:1 (v/v) mixture of diisopropyl ether/water and under stirring(600 mL). After 2 h of stirring, the heterogeneous mixture is allowed tostand overnight. The white precipitate is recovered by filtration,washed successively with a 1:1 (v/v) mixture of diisopropyl ether andwater (200 mL), then with water (100 mL). The solid obtained issolubilized in water (450 mL) by adjusting the pH to 7 by addition of anaqueous sodium hydroxide solution 10 N, then an aqueous sodium hydroxidesolution 1 N. The mixture is filtered through a 0.45 μm filter, thenpurified by ultrafiltration against a solution of NaCl 0.9%, then wateruntil the conductimetry of the permeate is less than 50 μS/cm. Thesolution of co-polyamino acid is then concentrated to approximately 30g/L theoretical and the pH is adjusted to 7.0. The aqueous solution isfiltered through a 0.2 μm filter and stored at 4° C.

-   Dry extract: 22.3 mg/g.-   DP (estimated by ¹H NMR)=29, thus i=0.034.-   The calculated average molecular weight of co-polyamino acid BB14 is    5089 g/mol.-   HPLC-aqueous SEC (calibrant PEG): Mn=4020 g/mol.

EXAMPLE BB15 Co-Polyamino Acid BB15—Sodium poly-L-glutamate Modified atOne of its Ends by Molecule BA3 and Having a Number Average MolecularWeight (Mn) of 3389 g/mol

By a method similar to the one used for the preparation of co-polyaminoacid BB14 applied to the hydrochloride salt of molecule BA3 (3.62 g,4.32 mmol) and to 25.0 g (94.97 mmol) of γ-benzyl-L-glutamateN-carboxyanhydride, a sodium poly-L-glutamate modified at one of itsends by molecule BA3 is obtained.

-   Dry extract: 30.4 mg/g.-   DP (estimated by ¹H NMR)=24, thus i=0.042.-   The calculated average molecular weight of co-polyamino acid BB15 is    4390 g/mol.-   HPLC-aqueous SEC (calibrant PEG): Mn=3389 g/mol.

EXAMPLE BB16 Co-Polyamino Acid BB16—Sodium poly-L-glutamate Modified atOne of its Ends by Molecule BA4 and Having a Number Average MolecularWeight (Mn) of 3300 g/mol

By a method similar to the one used for the preparation of co-polyaminoacid BB14 applied to the hydrochloride salt of molecule BA4 (5.70 g,5.70 mmol) and to 29.99 g (113.9 mmol) of γ-benzyl-L-glutamateN-carboxyanhydride, a sodium poly-L-glutamate modified at one of itsends by molecule BA4 is obtained.

-   Dry extract: 32.3 mg/g.-   DP (estimated by ¹H NMR)=23, thus i=0.043.-   The calculated average molecular weight of co-polyamino acid BB16 is    4399 g/mol.-   HPLC-aqueous SEC (calibrant PEG): Mn=3300 g/mol.

EXAMPLE BB17 Co-Polyamino Acid BB17—Sodium poly-L-glutamate Modified atOne of its Ends by Molecule BA3 and Having a Number Average MolecularWeight of 10,700 g/mol

By a method similar to the one used for the preparation of co-polyaminoacid BB14 applied to the hydrochloride salt of molecule BA3 (2.51 g, 3mmol) and to 52.7 g (200 mmol) of γ-benzyl-L-glutamateN-carboxyanhydride, a sodium poly-L-glutamate modified at one of itsends by molecule BA3 is obtained.

-   Dry extract: 24.5 mg/g.-   DP (estimated by ¹H NMR)=65, thus i=0.015.-   The calculated average molecular weight of co-polyamino acid BB17 is    10,585 g/g/mol.-   HPLC-aqueous SEC (calibrant PEG): Mn=10,700 g/g/mol.

EXAMPLE BB18 Co-Polyamino Acid BB18—Sodium poly-L-glutamate Modified atOne of its Ends by Molecule BA3 and Having a Number Average MolecularWeight of 6600 g/mol

By a method similar to the one used for the preparation of co-polyaminoacid BB14 applied to the hydrochloride salt of molecule BA3 (2.51 g, 3mmol) and to 31.6 g (120 mmol) of γ-benzyl-L-glutamateN-carboxyanhydride, a sodium poly-L-glutamate modified at one of itsends by molecule BA3 is obtained.

-   Dry extract: 27.3 mg/g.-   DP (estimated by ¹H NMR)=40, thus i=0.025.-   The calculated average molecular weight of co-polyamino acid BB18 is    6889 g/g/mol.-   HPLC-aqueous SEC (calibrant PEG): Mn=6600 g/g/mol.

EXAMPLE BB19 Co-Polyamino Acid BB19—Sodium poly-L-glutamate and Modifiedby Molecule BA3 and Having a Number Average Molecular Weight (Mn) of7700 g/mol

By a method similar to the one used for the preparation of co-polyaminoacid AB23 applied to the hydrochloride salt of molecule BA3 and toco-polyamino acid AB23-1, a sodium poly-L-glutamate modified by moleculeBA3 is obtained.

-   Dry extract: 25.3 mg/g.-   DP (estimated based on ¹H NMR): 60.-   Based on ¹H NMR: i=0.045.-   The calculated average molecular weight of co-polyamino acid BB19 is    11,188 g/g/mol.-   HPLC-organic SEC (calibrant PEG): Mn=7700 g/mol.

EXAMPLE BB20 Co-Polyamino Acid BB20—Sodium poly-L-glutamate Modified atOne of its Ends by Molecule BA5 and Having a Number Average MolecularWeight (Mn) of 2800 g/mol

By a method similar to the one used for the preparation of co-polyaminoacid BB14 applied to molecule BA5 in the form of a free amine (1.70 g,1.98 mmol) and to γ-benzyl-L-glutamate N-carboxyanhydride (11.46 g, 43.5mmol), a sodium poly-L-glutamate modified at one of its ends by moleculeBA5 is obtained.

-   Dry extract: 20.7 mg/g.-   DP (estimated by ¹H NMR)=23, thus i=0.043.-   The calculated average molecular weight of co-polyamino acid BB20 is    4295 g/g/mol.-   HPLC-aqueous SEC (calibrant PEG): Mn=2800 g/g/mol.

EXAMPLE BB21 Co-Polyamino Acid BB21—Sodium poly-L-glutamate Modified atOne of its Ends by Molecule BA3 and Having a Number Average MolecularWeight (Mn) of 1100 g/mol

By a method similar to the one used for the preparation of co-polyaminoacid BB14 applied to molecule BA3 in the form of a free amine (3.814 g,4.75 mmol) and to γ-benzyl-L-glutamate N-carboxyanhydride (10.0 g, 38.0mmol), a sodium poly-L-glutamate modified at one of its ends by moleculeBA3 is obtained.

-   Dry extract: 16.1 mg/g.-   DP (estimated by ¹H NMR)=9, thus i=0.11.-   The calculated average molecular weight of co-polyamino acid BB21 is    2123 g/g/mol.-   HPLC-aqueous SEC (calibrant PEG): Mn=1100 g/g/mol.

EXAMPLE BB22 Co-Polyamino Acid BB22—Sodium poly-L-glutamate Modified atOne of its Ends by Molecule BA6 and Having a Number Average MolecularWeight (Mn) of 3300 g/mol

By a method similar to the one used for the preparation of co-polyaminoacid BB14 applied to molecule BA6 in the form of a free amine (4.45 g,5.18 mmol) and to 30.0 g (113.96 mmol) of γ-benzyl-L-glutamateN-carboxyanhydride, a sodium poly-L-glutamate modified at one of itsends by molecule BA6 is obtained.

-   Dry extract: 29.0 mg/g.-   DP (estimated by ¹H NMR)=25, thus i=0.04.-   The calculated average molecular weight of co-polyamino acid BB22 is    4597 g/mol.-   HPLC-aqueous SEC (calibrant PEG): Mn=3300 g/mol.

EXAMPLE BB23 Co-Polyamino Acid BB23—Sodium poly-L-glutamate Modified atOne of its Ends by Molecule BA7 and Having a Number Average MolecularWeight (Mn) of 2900 g/mol

By a method similar to the one used for the preparation of co-polyaminoacid BB14 applied to molecule BA7 in the form of a free amine (3.05 g,4.01 mmol) and to γ-benzyl-L-glutamate N-carboxyanhydride (22.78 g, 86.5mmol), a sodium poly-L-glutamate modified at one of its ends by moleculeBA7 is obtained.

-   Dry extract: 16.9 mg/g.-   DP (estimated by ¹H NMR)=21, thus i=0.048.-   The calculated average molecular weight of co-polyamino acid BB23 is    3894 g/mol.-   HPLC-aqueous SEC (calibrant PEG): Mn=2900 g/mol.

EXAMPLE BB24 Co-Polyamino Acid BB27—Sodium poly-L-glutamate Modified atOne of its Ends by Molecule BA3 and Modified by Molecule BA3 and Havinga Number Average Molecular Weight (Mn) of 2300 g/mol

Co-polyamino acid BB24-1: poly-L-glutamic acid modified at one of itsends by molecule BA3 and capped at the other end by pidolic acid.

In a round-bottom flask dried in the oven, γ-benzyl-L-glutamateN-carboxyanhydride (122.58 g, 466 mmol) is placed under a vacuum for 30min, then anhydrous DMF (220 mL) is introduced. The mixture is stirredunder argon until the solubilization is complete, cooled to −10° C.,then a solution of molecule BA3 in the form of a free amine (17.08 g,21.3 mmol) in chloroform (40 mL) is introduced rapidly. The mixture isstirred between 0° C. and ambient temperature for 2 days, then heated at65° C. for 4 h. The reaction mixture is then cooled to 25° C., thenpidolic acid (13.66 g, 105.8 mmol) is added, HOBt (2.35 g, 15.3 mmol)and EDC (20.28 g, 105.8 mmol) are added. After 24 h of stirring at 25°C., the solution is concentrated under a vacuum to eliminate thechloroform and 50% of the DMF. The reaction mixture is then heated to55° C. and 1150 mL of methanol are introduced in 1 h. The reactionmixture is then cooled to 0° C. After 18 h, the white precipitate isrecovered by filtration, washed three times with 270 mL of diisopropylether, then dried under a vacuum at 30° C. to obtain a white solid. Thesolid is diluted in TFA (390 mL), and a solution of hydrobromic acid(HBr) at 33% in acetic acid (271 mL, 1547 mmol) is then added dropwiseand at 0° C. The solution is stirred for 2 h at ambient temperature,then poured dropwise into a 1:1 (v/v) mixture of diisopropyl ether/waterand under stirring (970 mL). After 2 h of stirring, the heterogeneousmixture is allowed to stand overnight. The white precipitate isrecovered by filtration, washed successively with diisopropyl ether (380mL), then two times with water (380 mL mL). The solid obtained issolubilized in water (3.6 L) by adjusting the pH to 7 by addition of anaqueous sodium hydroxide solution 10 N, then an aqueous sodium hydroxidesolution 1 N. The mixture is filtered through a 0.45 μm filter, thenpurified by ultrafiltration against a solution of NaCl 0.9%, a solutionof NaOH 0.1 N, a solution of NaCl 0.9%, a phosphate buffer solution (150mM), a solution of NaCl 0.9%, then water until the conductimetry of thepermeate is less than 50 μS/cm. The solution of co-polyamino acid isthen concentrated to approximately 30 g/L theoretical, filtered througha 0.2 μm filter, then acidified to pH 2 under stirring by addition of asolution of HCl at 37%. The precipitate is then recovered by filtration,washed two times with water, then dried under a vacuum at 30° C. toobtain a white solid.

Co-Polyamino Acid BB24

By a method similar to the one used for the preparation of co-polyaminoacid BB2 applied to molecule BA3 in the form of a free amine (1.206 g,1.50 mmol) and to co-polyamino acid BB24-1 (5.5 g, 33.4 mmol), a sodiumpoly-L-glutamate modified at one of its ends by molecule BA3 andmodified by molecule BA3 is obtained.

-   Dry extract: 19.0 mg/g.-   DP (estimated based on ¹H NMR): 22.-   Based on ¹H NMR: i=0.089.-   The calculated average molecular weight of co-polyamino acid BB24 is    4826 g/mol.-   HPLC-aqueous SEC (calibrant PEG): Mn=2300 g/mol

EXAMPLE BB25 Co-Polyamino Acid BB25—Sodium poly-L-glutamate Modified atOne of its Ends by Molecule BA3 and at the Other End by Molecule B8 andHaving a Number Average Molecular Weight (Mn) of 2000 g/mol

DCC (0.257 g, 1.24 mmol) and NHS (0.143 g, 1.24 mmol) are introducedinto a solution of molecule B8 (0.946 g, 1.24 mmol) in DMF (8 mL). After16 h of stirring at ambient temperature, the solution is filtered to beused directly in the next reaction.

In a round-bottom flask dried in the oven, γ-benzyl-L-glutamateN-carboxyanhydride (6.0 g, 22.8 mmol) is placed under a vacuum for 30min, then anhydrous DMF (14 mL) is introduced. The mixture is thenstirred under argon until the dissolution is complete, cooled to 0° C.,then a solution of molecule BA3 in the form of a free amine (0.832 g,1.04 mmol) in chloroform (2.0 mL) is introduced rapidly. After 18 h ofstirring at 0° C., the previously prepared solution of molecule B8 isadded. The solution is stirred between 0° C. and ambient temperature for22 h, then poured dropwise into diisopropyl ether (0.34 L) understirring. The precipitate is recovered by filtration, washed withdiisopropyl ether (7 times 15 mL), then dried under a vacuum at 0° C. toyield a white solid. The solid is diluted in TFA (23 mL), then thesolution is cooled to 4° C. A solution of HBr at 33% in acetic acid (15mL, 85.7 mmol) is then added dropwise. The mixture is stirred at ambienttemperature for 2 h, then poured dropwise into a 1:1 (v/v) mixture ofdiisopropyl ether and water under stirring (0.28 L). After 2 h ofstirring, the heterogeneous mixture is allowed to stand overnight. Thewhite precipitate is recovered by filtration, washed two times with a1:1 (v/v) mixture of diisopropyl ether and water (24 mL), then two timeswith water (24 mL). The solid obtained is then solubilized in water(0.16 L) by adjusting the pH to 12 by addition of an aqueous sodiumhydroxide solution 10 N, then an aqueous sodium hydroxide solution 1 N.After 30 min, the pH is adjusted to 7 by slow addition of an aqueous HClsolution 1 N. The solution is filtered through a 0.45 μm filter, thenpurified by ultrafiltration against a solution of NaCl 0.9% %, thenwater until the conductimetry of the permeate is less than 50 μS/cm. Thesolution obtained is filtered through a 0.2 μm filter and stored at 2-8°C.

-   Dry extract: 18.9 mg/g.-   DP (estimated based on ¹H NMR): 22.-   Based on ¹H NMR: i=0.09.-   The calculated average molecular weight of co-polyamino acid BB25 is    4871 g/mol.-   HPLC-aqueous SEC (calibrant PEG): Mn=2000 g/mol

C. Compositions EXAMPLE CV1 Preparation of a Solution of Human Amylin at0.6 mg/mL Containing m-cresol (29 mM), glycerol (174 mM) at pH 7.4

A concentrated solution of human amylin at 3 mg/mL is prepared bydissolution of human amylin in the form of a powder purchased fromAmbioPharm. This solution is added to a concentrated solution ofexcipients (m-cresol, glycerol) in such a manner as to obtain theintended final composition. The final pH is adjusted to 7.4 by additionof NaOH/HCl.

EXAMPLE CV2 Preparation of a Solution of Human Amylin at 0.6 mg/mLContaining Co-Polyamino Acid BB15, m-cresol (29 mM) and glycerol (174mM) at pH 7.4

A concentrated solution of co-polyamino acid BB15 and excipients isprepared by adding concentrated solutions of excipients (m-cresol,glycerol) to a concentrated solution of co-polyamino acid BB15.

The concentrated solution of human amylin at 3 mg/mL C1 is added to thisconcentrated solution of co-polyamino acid BB15 and excipients in such amanner as to obtain the final compositions CV5 to CV11 (table 1g). Thefinal pH is adjusted to 7.4 by addition of NaOH/HCl.

TABLE 1g Compositions and visual appearance of solutions of human amylinat pH 7.4 at different concentrations of co-polyamino acid BB15. RatioBB15/human Concentration of co- amylin polyamino acid BB15 Visualappearance Solution mol/mol mg/mL mM of the solution CV1 — — — Clear CV55 3 0.73 Clear CV6 6 3.6 0.88 Clear CV7 7 4.2 1.03 Clear CV8 8 4.8 1.17Clear CV9 9 5.4 1.32 Clear CV10 10 6 1.47 Clear CV11 17 10.5 2.57 Clear

EXAMPLE CY1 Preparation of a Solution of pramlintide at 0.9 mg/mLContaining m-cresol (29 mM) and glycerol (174 mM) at pH 7.4

A concentrated solution of pramlintide at 5 mg/mL is prepared bydissolution of pramlintide in the form of a powder purchased from Hybio.This solution is added to a concentrated solution of excipients(m-cresol, glycerol) in such a manner as to obtain the intended finalcomposition. The final pH is adjusted to 7.4 by addition of NaOH/HCl.

EXAMPLE CW1 Preparation of a Solution of pramlintide at 0.4 mg/mLContaining phenol (30 mM), glycerol (174 mM) and glycylglycine (8 mM) atpH 7.4

By a method similar to the one used in Example CY1, a solution ofpramlintide at 0.4 mg/mL containing phenol (30 mM), glycerol (174 mM)and glycylglycine (8 mM) at pH 7.4 is obtained.

EXAMPLE CY0 Preparation of a Solution of pramlintide at 0.9 mg/mLContaining Co-Polyamino Acid BB15, m-cresol (29 mM) and glycerol (174mM) at pH 7.4

A concentrated solution of co-polyamino acid BB15 and excipients isprepared by adding concentrated solutions of excipients (m-cresol,glycerol) to a concentrated solution of co-polyamino acid BB15.

A concentrated solution of pramlintide at 5 mg/mL is added to thisconcentrated solution of co-polyamino acid BB15 and excipients in such amanner as to obtain the final compositions CY2 to CY7 (table 3). Thefinal pH is adjusted to 7.4 by addition of NaOH/HCl.

TABLE 3 Compositions and visual appearance of solutions of pramlintideat pH 7.4 at different concentrations of co-polyamino acids BB15. RatioConcentration of co- BB15/pramlintide polyamino acid BB15 Visualappearance Solution mol/mol mg/mL mM of the solution CY1 — — — Clear CY22 1.8 0.44 Clear CY3 3 2.7 0.66 Clear CY4 4 3.6 0.88 Clear CY5 5 4.51.10 Clear CY6 6 5.4 1.32 Clear CY7 10 9 2.20 Clear

EXAMPLE CW0 Preparation a Solution of pramlintide at 0.4 mg/mLContaining Co-Polyamino Acid BB15, phenol (30 mM), glycerol (174 mM) andglycylglycine (8 mM) at pH 7.4

By a method similar to the one used in Example CY0, starting with asolution of pramlintide at 0.4 mg/mL CW1, solutions of pramlintide at0.4 mg/mL containing co-polyamino acid BB15, phenol (30 mM), glycerol(174 mM) and glycylglycine (8 mM) at pH 7.4, CW2 and CW3 are obtained.

TABLE 4 Compositions and visual appearance of solutions of pramlintideat 0.4 mg/mL at pH 7.4 at different concentrations of co-polyamino acidBB15. Concentration of co- Ratio polyamino acid BB15 BB15/pramlintideVisual appearance Solution mg/mL mM mol/mol of the solution CW2 2.4 0.596 Clear CW3 4 0.98 10 Clear

EXAMPLE CP0 Preparation of a Solution of pramlintide at 0.9 mg/mLContaining Different Co-Polyamino Acids of the Invention, m-cresol (29mM) and glycerol (174 mM) at pH 7.4

By a method similar to the one described in Example CY0, solutions ofpramlintide at 0.9 mg/mL containing different co-polyamino acids of theinvention, m-cresol (29 mM) and glycerol (174 mM) at pH 7.4, CP2 to CP12are obtained.

TABLE 8 Compositions and visual appearance of solutions of pramlintideat 0.9 mg/mL at pH 7.4 in the presence of different co-polyamino acids.Ratio co- Concentration polyamino of co- acid/ Visual Co- polyamino acidpramlintide appearance of Solution polyaminoacid mg/mL mM mol/mol thesolution CP2 BB15 5 1.22 5.4 Clear 10 2.45 10.8 Clear CP3 BB14 5 0.984.3 Clear 10 1.96 8.6 Clear CP4 AB17 5 1.11 4.9 Clear 10 1.22 9.8 ClearCP5 AB15 5 0.99 4.4 Clear 10 1.99 8.8 Clear CP6 AB14 5 1.48 6.5 Clear 102.96 13 Clear CP10 BB18 5 0.72 3.2 Clear 10 1.44 6.3 Clear CP11 BB9 5 14.4 Clear 10 2.01 8.8 Clear CP12 BB2 5 1.27 5.6 Clear 10 2.54 11.2 Clear

EXAMPLE CH1 Preparation of a Solution of pramlintide at 0.6 mg/mLContaining m-cresol (29 mM) and glycerol (174 mM) at pH 6.6

A concentrated solution of pramlintide at 5 mg/mL is prepared bydissolution of pramlintide in the form of a powder purchased fromAmbiopharm. This solution is added to a concentrated solution ofexcipients (m-cresol, glycerol) in such a manner as to obtain theintended final composition. The final pH is adjusted to 6.6 by additionof NaOH/HCl.

EXAMPLE CH0 Preparation of a Solution of pramlintide at 0.6 mg/mLContaining Co-Polyamino Acid BB15, m-cresol (29 mM) and glycerol (174mM) at pH 6.6

A concentrated solution of copolyamino acid BB15 and excipients isprepared by adding concentrated solutions of excipients (m-cresol,glycerol) to a concentrated solution of co-polyamino acid BB15.

A concentrated solution of pramlintide at 5 mg/mL at pH 4 is added tothis concentrated solution of co-polyamino acid BB15 and excipients insuch a manner as to obtain the final compositions CH2 to CH8 (table 9).The final pH is adjusted to 6.6 by addition of NaOH/HCl.

TABLE 9 Compositions and visual appearance of solutions of pramlintideat pH 6.6 at different concentrations of co-polyamino acid BB15. Ratio/Concentration of co- BB15 pramlintide polyamino acid BB15 Visualappearance Solution mol/mol mg/mL mM of the solution CH1 — — — Clear CH22 1.3 0.29 Clear CH3 3 2 0.45 Clear CH4 4 2.7 0.61 Clear CH5 6 4 0.90Clear CH6 8 5.3 1.19 Clear CH7 10 6.7 1.50 Clear CH8 15 10 2.24 Clear

EXAMPLE CI0 Preparation of a Solution of pramlintide at 0.6 mg/mLContaining Different Co-Polyamino Acids of the Invention, m-cresol (29mM) and glycerol (174 mM) at pH 6.6

By a method similar to the one described in example CH0, solutions ofpramlintide at 0.6 mg/mL containing different co-polyamino acids of theinvention, m-cresol (29 mM) and glycerol (174 mM) at pH 6.6, CI1 to CI14are obtained.

TABLE 10 Compositions and visual appearance of solutions of pramlintideat pH 6.6 in the presence of different co-polyamino acids. Ratioco-polyamino Visual Co- Concentration of acid/ appearance polyaminoco-polyamino acid pramlintide of Solution acid mg/mL mM mol/mol thesolution CI1 BB20 1.3 0.3 2 Clear 2.6 0.6 4 Clear CI2 BB21 1.3 0.6 4Clear CI3 AB22 2.4 0.3 2 Clear CI4 BB24 2.9 0.6 4 Clear CI5 BB25 1.5 0.32 Clear 3 0.6 4 Clear CI6 AB23 3.4 0.23 2 Clear CI7 AB28 2.3 0.3 2 Clear4.7 0.6 4 Clear CI8 AB24 1.2 0.15 1 Clear 2.4 0.3 2 Clear CI9 AB25 1.30.15 1 Clear 2.6 0.3 2 Clear CI10 AB26 0.7 0.15 1 Clear 1.5 0.3 2 ClearCI11 AB27 1.3 0.15 1 Clear 2.7 0.3 2 Clear CI12 AB31 1.3 0.15 1 Clear2.5 0.3 2 Clear CI13 AB29 8.9 1.15 7.6 Clear CI14 AB32 1.3 0.15 1 Clear2.5 0.3 2 Clear

EXAMPLE CT0 Preparation of a Solution of pramlintide at 0.6 mg/mLContaining Co-Polyamino Acid AB14, m-cresol (29 mM), glycerol (174 mM),NaCl and zinc chloride at pH 6.6

A concentrated solution of co-polyamino acid AB14 and excipients isprepared by adding concentrated solutions of excipients (m-cresol,glycerol, NaCl, zinc chloride) to a concentrated solution ofco-polyamino acid AB14.

A concentrated solution of pramlintide at 5 mg/mL at pH 4 is added tothis concentrated solution of co-polyamino acid AB14 and excipients insuch a manner as to obtain the final composition CT1 to CT5 (table 11).The final pH is adjusted to 6.6 by addition of NaOH/HCl.

TABLE 11 Compositions and visual appearance of the solutions ofpramlintide at pH 6.6 in the presence of co-polyamino acid AB14 and ofdifferent contents of sodium chloride and zinc chloride. Concentrationof Visual Co- co-polyamino appearance polyamino acid [NaCl] [ZnCl₂] ofthe Solution acid mg/mL mM (mM) (mM) solution CT1 AB14 6.3 1.87 — 0.75Clear CT2 AB14 6.3 1.87 50 — Clear CT3 AB14 6.3 1.87 100 — Clear CT4AB14 6.3 1.87 50 0.75 Clear CT5 AB14 6.3 1.87 100 0.75 Clear

EXAMPLE CS0 Preparation of a Solution of pramlintide at 0.6 mg/mLContaining Different Co-Polyamino Acids of the Invention, m-cresol (29mM), glycerol (174 mM), NaCl and zinc chloride at pH 6.6

By a method similar to the one described in example CA4, solutions ofpramlintide at 0.6 mg/mL containing different co-polyamino acids of theinvention, m-cresol (29 mM) and glycerol (174 mM), sodium chloride andzinc chloride at pH 6.6, BS1 to BS11 are obtained.

TABLE 12 Compositions and visual appearance of the solutions ofpramlintide at pH 6.6 in the presence of different co-polyamino acidsand of different contents of sodium chloride and zinc chlorideConcentration of Visual Co- co-polyamino appearance polyamino acid[NaCl] [ZnCl₂] of the Solution acid mg/mL mM (mM) (mM) solution CS1 AB157.8 1.6 — — Clear CS2 AB15 11.7 2.3 — — Clear CS3 AB15 3.9 0.8 50 —Clear CS4 AB15 6.3 1.3 50 — Clear CS5 AB15 7.8 1.6 50 — Clear CS6 AB153.9 0.8 100 — Clear CS7 AB16 12.4 1.5 — — Clear CS8 AB16 16.7 2.1 — —Clear CS9 AB16 7.4 0.9 50 — Clear CS10 AB16 12.4 1.5 50 — Clear CS11AB16 7.4 0.9 50 1 Clear

EXAMPLE CX1 Preparation of a Solution of Human Amylin at 0.6 mg/mL andHuman Insulin at 100 IU/mL Containing m-cresol (29 mM), glycerol (174mM) and zinc chloride (229 μM) at pH 7.4

The concentrated solution of human amylin at 3 mg/mL CV1 is added to aconcentrated solution of excipients (m-cresol, glycerol). A solution ofhuman insulin at 500 IU/mL is prepared by dissolution of human insulinin the form of a powder purchased from Amphastar. This solution is addedto the concentrated solution of human amylin and excipients in such amanner as to obtain the intended final composition. The final pH isadjusted to 7.4 by addition of NaOH/HCl.

EXAMPLE CX2 Preparation of a Solution of Human Amylin at 0.6 mg/mL andof Human Insulin at 100 IU/mL Containing Co-Polyamino acid BB15,m-cresol (29 mM), glycerol (174 mM) and zinc chloride (229 μM) at pH 7.4

A concentrated solution of co-polyamino acid BB15 and excipients isprepared by adding concentrated solutions of excipients (m-cresol,glycerol, zinc chloride) to a concentrated solution of co-polyamino acidBB15.

A concentrated solution of human amylin at 3 mg/mL, then a solution ofhuman insulin at 500 IU/mL are added to the concentrated solution ofco-polyamino acid BB15 and excipients in such a manner as to obtain theintended final composition (table 13). The final pH is adjusted to 7.4by addition of NaOH/HCl.

The solutions CX1, CX6, CX10 and CX11 are prepared according to theabove protocol.

TABLE 11 Composition and visual appearance of solutions of human amylinand of human insulin at pH 7.4 at different concentrations ofco-polyamino acid BB15. Ratio BB15/human Concentration of co- amylinpolyamino acid BB15 Visual appearance Solution mol/mol mg/mL mM of thesolution CX1 — — — Turbid CX6 6 3.6 0.88 Clear CX10 10 6 1.47 Clear CX1117 10.5 2.57 Clear

In the presence of co-polyamino acid BB15, a clear solution of humanamylin (0.6 mg/mL) and of human insulin (100 IU/mL) is obtained at pH7.4.

EXAMPLE CN1 Preparation of a Solution of pramlintide at 0.4 mg/mL and ofHuman Insulin at 100 IU/mL Containing phenol (30 mM), glycerol (174 mM),glycylglycine (8 mM) and zinc chloride (229 μM) at pH 7.4

A concentrated solution of pramlintide at 5 mg/mL is added to aconcentrated solution of excipients (m-cresol, glycerol, glycylglycine,zinc chloride). A solution of human insulin at 500 IU/mL is added tothis concentrated solution of pramlintide and excipients in such amanner as to obtain the intended final composition. The final pH isadjusted to 7.4 by addition of NaOH/HCl.

EXAMPLE CR1 Preparation of a Solution of pramlintide at 0.9 mg/mL and ofHuman Insulin at 100 IU/mL Containing m-cresol (29 mM), glycerol (174mM) and zinc chloride (229 μM) at pH 7.4

By a method similar to the one used in example CN1, a solution ofpramlintide at 0.9 mg/mL and of human insulin at 100 IU/mL containingm-cresol (29 mM), glycerol (174 mM) and zinc chloride (229 μM) at pH 7.4is obtained.

EXAMPLE CN0 Preparation of a Solution of pramlintide at 0.4 mg/mL and ofHuman Insulin at 100 IU/mL Containing Co-Polyamino Acid BB15, phenol (30mM), glycerol (174 mM), glycylglycine (8 mM) and zinc chloride (229 μM)at pH 7.4

A concentrated solution of co-polyamino acid BB15 and excipients isprepared by adding concentrated solutions of excipients (m-cresol,glycerol, glycylglycine, zinc chloride) to a concentrated solution ofco-polyamino acid BB15.

A concentrated solution of pramlintide at 5 mg/mL, then a solution ofhuman insulin at 500 IU/mL are added to this concentrated solution ofco-polyamino acid BB15 and excipients in such a manner as to obtain theintended final composition (table 14). The final pH is adjusted to 7.4by addition of NaOH/HCl.

The solutions CN2 and CN3 are prepared according to the above protocol.

TABLE 14 Compositions and visual appearance of the solutions ofpramlintide at 0.4 mg/mL and of human insulin at 100 IU/mL at pH 7.4 atdifferent concentrations of co-polyamino acid BB15. Concentration ofco-polyamino Ratio acid BB15 BB15/pramlintide Visual appearance Solutionmg/mL mM mol/mol of the mixture CN1 — — Turbid CN2 2.4 0.59 6 Clear CN34 0.98 10 Clear

In the presence of co-polyamino acid BB15, a clear solution ofpramlintide (0.4 mg/mL) and of human insulin (100 IU/mL) is obtained atpH 7.4.

EXAMPLE CR0 Preparation of a Solution of pramlintide at 0.9 mg/mL and ofHuman Insulin at 100 IU/mL Containing Co-Polyamino acid BB15, m-cresol(29 mM), glycerol (174 mM) and zinc chloride (229 μM) at pH 7.4

By a method similar to the one used in example CN0, a solution ofpramlintide at 0.9 mg/mL and of human insulin at 100 IU/mL containingco-polyamino acid BB15, m-cresol (29 mM), glycerol (174 mM) and zincchloride (229 μM) at pH 7.4 is obtained.

The solutions CR2 to CR4 and CU2 to CU8 are prepared according to theabove protocol.

TABLE 15 Compositions and appearance of the solutions of pramlintide at0.9 mg/mL and of human insulin at 100 IU/mL at pH 7.4 at differentconcentrations of co-polyamino acid BB15. Concentration of co-polyaminoacid Ratio BB15 BB15/pramlintide Visual appearance of Solution mg/mL mMmol/mol the solution CR1 — — Turbid CR2 2.7 0.66 3 Clear CR3 3.6 0.88 4Clear CR4 4.5 1.10 5 Clear CU2 0.9 0.22 1 Clear CU3 1.8 0.44 2 Clear CU75.4 1.32 6 Clear CU8 9 2.20 10 Clear

In the presence of co-polyamino acid BB15, a clear solution ofpramlintide (0.9 mg/mL) and of human insulin (100 IU/mL) at pH 7.4 isobtained.

EXAMPLE CG0 Preparation of a Solution of pramlintide at 0.9 mg/mL and ofHuman Insulin at 100 IU/mL Containing Different Co-Polyamino Acids ofthe Invention, m-cresol (29 mM), glycerol (174 mM) and zinc chloride(229 μM) at pH 7.4

By a method similar to example CN0, a solution of pramlintide at 0.9mg/mL and of human insulin at 100 IU/mL containing a co-polyamino acidof the invention, m-cresol (29 mM), glycerol (174 mM) and zinc chloride(229 μM) at pH 7.4 is obtained.

The solutions CG2 to CG12 are prepared according to the above-describedprotocol.

TABLE 16 Compositions and visual appearance of solutions of pramlintideat 0.9 mg/mL and of human insulin at 100 IU/mL at pH 7.4 at differentconcentrations of co-polyamino acids. Ratio Co-polyamino Visual Co-Concentration of acid/ appearance polyamino co-polyamino acidpramlintide of Solution acid mg/mL mM mol/mol the solution CG2 BB15 51.22 5.4 Clear 10 2.45 10.8 Clear CG3 BB14 5 0.98 4.3 — 10 1.96 8.6Clear CG4 AB17 5 1.11 4.9 Clear 10 1.22 9.8 Clear CG5 AB15 5 0.99 4.4 —10 1.99 8.8 Clear CG6 AB14 5 1.48 6.5 — 10 2.96 13 Clear CG10 BB18 50.72 3.2 Clear 10 1.44 6.3 Clear CG11 BB9 5 1 4.4 Clear 10 2.01 8.8Clear CG12 BB2 5 1.27 5.6 Clear 10 2.54 11.2 Clear

EXAMPLE CD1 Preparation of a Solution of pramlintide at 0.9 mg/mL and ofInsulin Lispro at 100 IU/mL Containing m-cresol (29 mM), glycerol (174mM) and zinc chloride (300 μM) at pH 7.4

A concentrated solution of pramlintide at 5 mg/mL is added to aconcentrated solution of excipients (m-cresol, glycerol, zinc chloride).A solution of insulin lispro at 500 IU/mL is added to this concentratedsolution of pramlintide and excipients in such a manner as to obtain theintended final composition. The final pH is adjusted to 7.4 by additionof NaOH/HCl.

EXAMPLE CD0 Preparation of a Solution of pramlintide at 0.9 mg/mL and ofInsulin Lispro at 100 IU/mL Containing Co-Polyamino Acid BB15, m-cresol(29 mM), glycerol (174 mM) and zinc chloride (300 μM) at pH 7.4

A concentrated solution of co-polyamino acid BB15 and excipients isprepared by adding concentrated solutions of excipients (m-cresol,glycerol, zinc chloride) to a concentrated solution of co-polyamino acidBB15.

A concentrated solution of pramlintide at 5 mg/mL, then a solution ofinsulin lispro at 500 IU/mL are added to the concentrated solution ofco-polyamino acid BB15 and excipients in such a manner as to obtain theintended final composition. The final pH is adjusted to pH 7.4 byaddition of NaOH/HCl.

The solutions CD3 to CD9 are prepared according to the above-describedprotocol.

TABLE 17 Compositions and visual appearance of the solutions ofpramlintide at 0.9 mg/mL and of insulin lispro at 100 IU/mL at pH 7.4 atdifferent concentrations of co-polyamino acid BB15. Ratio Concentrationof co- BB15/pramlintide polyamino acid BB15 Visual appearance Solutionmol/mol mg/ml mM of the solution CD1 — — — Turbid CD3 2 1.8 0.44 ClearCD4 3 2.7 0.66 Clear CD5 4 3.6 0.88 Clear CD6 5 4.5 1.10 Clear CD7 6 5.41.32 Clear CD8 10 9 2.20 Clear CD9 15 13.5 3.30 Clear

In the presence of co-polyamino acid BB15, a clear solution ofpramlintide (0.9 mg/mL) and of insulin lispro (100 IU/mL) at pH 7.4 isobtained.

EXAMPLE CK1 Preparation of a Solution of pramlintide at 0.6 mg/mL and ofHuman Insulin at 100 IU/mL Containing m-cresol (29 mM), glycerol (174mM) and zinc chloride (229 μM) at pH 6.6

By a method similar to the one used in example BR1, a solution ofpramlintide at 0.6 mg/mL and of human insulin at 100 IU/mL containingm-cresol (29 mM), glycerol (174 mM) and zinc chloride (229 μM) at pH 6.6is obtained.

EXAMPLE CK0 Preparation of a Solution of pramlintide at 0.6 mg/mL and ofHuman Insulin at 100 IU/mL Containing Co-Polyamino Acid BB15, m-cresol(29 mM), glycerol (174 mM) and zinc chloride (229 μM) at pH 6.6

By a method similar to the one used in example BR0, a solution ofpramlintide at 0.6 mg/mL and of human insulin at 100 IU/mL containingco-polyamino acid BB15, m-cresol (29 mM), glycerol (174 mM) and zincchloride (229 μM) at pH 6.6 is obtained.

The solutions CK2 to CK8 are prepared according to the above protocol.

TABLE 18 Compositions and visual appearance of the solutions ofpramlintide at 0.6 mg/mL and of human insulin at 100 IU/mL at pH 6.6 atdifferent concentrations of co-polyamino acid BB15. Ratio Concentrationof co- Visual BB15/pramlintide polyamino acid BB15 appearance ofSolution mol/mol mg/mL mM the solution CK1 — — — Turbid CK2 2 1.3 0.29Clear CK3 3 2 0.45 Clear CK4 4 2.7 0.61 Clear CK5 6 4 0.90 Clear CK6 85.3 1.19 Clear CK7 10 6.7 1.50 Clear CK8 15 10 2.24 Clear

In the presence of co-polyamino acid BB15, a clear solution ofpramlintide (0.6 mg/mL) and of human insulin (100 IU/mL) at pH 6.6 isobtained.

EXAMPLE CF1 Preparation of Compositions Containing VariableConcentrations of pramlintide, of Human Insulin at 100 IU/mL, m-cresol(29 mM), glycerol (174 mM) and zinc chloride (229 μM) at pH 6.6

By a method similar to the one used in example CR1, solutions containingdifferent concentrations of pramlintide, of human insulin at 100 IU/mL,m-cresol (29 mM), glycerol (174 mM) and zinc chloride (229 μM) at pH 6.6are obtained.

TABLE 18a Compositions and visual appearance of the solutions ofpramlintide at different concentrations and of human insulin at 100IU/mL at pH 6.6. Concentration of pramlintide Visual appearance of theSolution (mg/mL) solution CF1A 0.9 turbid CF1B 0.8 turbid CF1C 0.6turbid CF1D 0.3 turbid CF1E 0.2 turbid

EXAMPLE CF0 Preparation of Compositions Containing VariableConcentrations of pramlintide and of Human Insulin at 100 IU/mL in thePresence of Co-Polyamino acid AB24, m-cresol (29 mM), glycerol (174 mM)and zinc chloride (229 μM) at pH 6.6

By a method similar to the one used in example CR0, solutions containingdifferent concentrations of pramlintide, of human insulin at 100 IU/mL,co-polyamino acid AB24, m-cresol (29 mM), glycerol (174 mM) and zincchloride (229 μM) at pH 6.6 are obtained.

TABLE 18b Compositions and visual appearance of the solutions ofpramlintide at different concentrations and of human insulin at 100IU/mL in the presence of co-polyamino acid AB24 at pH 6.6. Ratio co-Visual Concentration of polyamino ap- Concentration co-polyamino acidacid/ pearance So- of pramlintide AB24 pramlintide of the lution (mg/mL)mg/mL mM mol/mol solution CF2 0.9 5.4 0.67 3 clear CF3 0.8 4.8 0.6 3clear CF4 0.6 3.6 0.45 3 clear CF5 0.3 1.8 0.22 3 clear CF6 0.2 1 0.1252.5 clear

EXAMPLE CM0 Preparation of a Solution of pramlintide at 0.6 mg/mL and ofHuman Insulin at 100 IU/mL Containing Different Co-Polyamino Acids ofthe Invention, m-cresol (29 mM), glycerol (174 mM) and zinc chloride(229 μM) at pH 6.6

By a method similar to example CG0, solutions of pramlintide at 0.6mg/mL and of human insulin at 100 IU/mL containing differentco-polyamino acids of the invention, m-cresol (29 mM), glycerol (174 mM)and zinc chloride (229 μM) at pH 6.6 are obtained.

The solutions CM1 to CM18 are prepared according to the above-describedprotocol.

TABLE 19 Compositions and visual appearance of the solutions ofpramlintide at 0.6 mg/mL and of human insulin at 100 IU/mL at pH 6.6 inthe presence of different co-polyamino acids. Ratio co-polyamino VisualCo- Concentration of acid/ appearance polyamino co-polyamino acidpramlintide of Solution acid mg/mL mM mol/mol the solution CM1 BB20 2.60.61 4 Clear 5.3 1.22 8 Clear CM2 BB21 1.3 0.6 4 Clear CM3 AB22 2.4 0.32 Clear CM4 BB24 2.9 0.6 4 Clear CM5 BB23 3 0.76 5 Clear CM6 BB25 1.50.3 2 Clear CM7 BB22 2.7 0.6 4 Clear CM8 AB23 7.7 0.69 4.6 Clear CM9BB19 4.7 0.4 2.8 Clear CM10 AB28 2.3 0.3 2 Clear CM11 AB24 1.2 0.15 1Clear 2.4 0.3 2 Clear 3.6 0.45 3 CM12 AB25 2.6 0.3 2 Clear CM13 AB26 1.50.3 2 Clear 2.3 0.5 3 Clear CM14 AB27 1.3 0.15 1 Clear 2.7 0.3 2 ClearCM15 AB30 1.2 0.15 1 Clear 2.3 0.3 2 Clear CM16 AB31 1.3 0.15 1 Clear2.5 0.3 2 Clear CM17 AB29 5.9 0.8 5 Clear 8.9 1.15 7.6 Clear CM18 AB322.5 0.3 2 Clear

EXAMPLE CQ1 Preparation of a Solution of pramlintide at 0.6 mg/mL and ofHuman Insulin at 100 IU/mL at pH 6.6 Containing Co-Polyamino Acid AB14,m-cresol (29 mM), glycerol (174 mM), sodium chloride (100 mM) and zincchloride (1 mM)

A concentrated solution of co-polyamino acid AB14 and excipients isprepared by adding concentrated solutions of excipients (m-cresol,glycerol, sodium chloride, zinc chloride) to a concentrated solution ofco-polyamino acid AB14.

A concentrated solution of pramlintide at 5 mg/mL at pH 4, then asolution of human insulin at 500 IU/mL are added to this concentratedsolution of co-polyamino acid AB14 and excipients in such a manner as toobtain the intended final composition. The final pH is adjusted to 6.6by addition of NaOH/HCl.

The solution CQ1 is prepared according to the above protocol.

EXAMPLE CQ0 Preparation of a Solution of pramlintide at 0.6 mg/mL and ofHuman Insulin at 100 IU/mL Containing Different Co-Polyamino Acids ofthe Invention, m-cresol (29 mM), glycerol (174 mM) and DifferentContents of sodium chloride and of zinc chloride

By a method similar to example CQ0, solutions of pramlintide at 0.6mg/mL and of human insulin at 100 IU/mL containing differentco-polyamino acids of the invention, m-cresol (29 mM), glycerol (174mM), sodium chloride and zinc chloride at pH 6.6 are obtained.

The solutions CQ2 to CQ12 are prepared according to the above protocol.

TABLE 20 Compositions and visual appearance of the solutions ofpramlintide at 0.6 mg/mL and of human insulin at 100 IU/mL pH 6.6 in thepresence of different co-polyamino acids and of different contents ofsodium chloride and of zinc chloride. Concentration of Visual Co-co-polyamino appearance polyamino acid [NaCl] [ZnCl₂] of the Solutionacid mg/mL mM (mM) (mM) solution CQ1 AB14 6.3 1.87 100 1 Clear CQ2 AB157.8 1.6 — 0.23 Clear CQ3 AB15 11.7 2.3 — 0.23 Clear CQ4 AB15 3.9 0.8 500.23 Clear CQ5 AB15 6.3 1.3 50 0.23 Clear CQ6 AB15 7.8 1.6 50 0.23 ClearCQ7 AB15 3.9 0.8 100 0.23 Clear CQ8 AB15 6.3 1.3 100 0.23 Clear CQ9 AB157.8 1.6 100 0.23 Clear CQ10 AB16 7.4 0.9 50 0.23 Clear CQ11 AB16 12.41.5 50 0.23 Clear CQ12 AB16 7.4 0.9 50 1 Clear

EXAMPLE CZ0 Preparation of a Solution of pramlintide at 0.6 mg/mLContaining DMPG, m-cresol (29 mM), glycerol (174 mM) at p-H 6.6

A concentrated solution of DMPG and excipients is prepared by addingconcentrated solutions of excipients (m-cresol, glycerol) to aconcentrated solution of DMPG.

A concentrated solution of pramlintide at 10 mg/mL is added to thisconcentrated solution of DMPG and excipients in such a manner as toobtain the intended final composition. The final pH is adjusted to 6.6by addition of NaOH/HCl.

EXAMPLE CZ1 Preparation of a Solution of pramlintide at 0.6 mg/mLContaining DMPG (4.5 mM), phenol (30 mM), glycerol (174 mM) andglycylglycine (8 mM) at pH 7.4

TABLE 21 Compositions and visual appearance of the solutions ofpramlintide at 0.6 mg/mL in the presence of DMPG. Visual Concentrationappearance DMPG pramlintide of the Solution (mM) (mg/mL) pH Excipientssolution CZ0 4.5 0.6 6.6 m-cresol 29 mM Clear Glycerol 174 mM CZ1 4.50.6 7.4 phenol 30 mM Clear Glycylglycine 8 mM Glycerol 174 mM

EXAMPLE CA1 Preparation of a Solution of pramlintide at 1 mg/mLContaining m-cresol (20 mM), mannitol (43 mg/mL) and a sodium acetateBuffer, at pH 4

A concentrated solution of pramlintide at 10 mg/mL is added to aconcentrated solution of excipients (m-cresol, mannitol, sodium acetate)in such a manner as to obtain the intended final composition. The finalpH is adjusted to 4 by addition of NaOH/HCl. The clear solution isfiltered (0.22 μm) and introduced into 3 mL glass cartridges forinjector pen.

EXAMPLE CA2 Introduction into Cartridges of a Commercial Solution ofHuman Insulin at 100 IU/mL (Humulin®) Containing m-cresol (23 mM),glycerol (174 mM) and zinc chloride (230 μM)

A commercial solution of Humulin® in a 10 mL vial is collected andintroduced into 3 mL glass cartridges for injector pen.

EXAMPLE CA3 Preparation of a Solution of pramlintide at 0.6 mg/mL and ofHuman Insulin at 100 IU/mL Containing Co-Polyamino Acid BB15 (3.1mg/mL), m-cresol (29 mM), glycerol (35 mM), mannitol (2.6% % w/v), Tris(18.75 mM), sodium acetate (18 mM), and zinc chloride (260 μM) at pH 6.2

3 mL glass vials are filled with 0.5 mL of a solution containing 10mg/mL of co-polyamino acid BB15 and 60 mM of Tris at pH 8.3. Thesolution is lyophilized in such a manner as to obtain 3 mL vialscontaining 5 mg of co-polyamino acid BB15 and 30 μmol of Tris.

A solution of human insulin at 500 IU/mL containing 23 mM of m-cresol,174 mM of glycerol and 260 μM of zinc chloride at pH 7.4 is prepared byadding concentrated solutions of excipients (m-cresol, glycerol, zincchloride) to a concentrated solution of human insulin concentrated at760 IU/mL.

In the vial containing 5 mg of co-polyamino acid BB15 and 30 μmol ofTris, the following are introduced successively:

-   -   0.96 mL of the solution of Pramlintide at 1 mg/mL at pH 4        described in example CA1;    -   0.32 mL of sterile water for injection;    -   0.32 mL of concentrated human insulin at 500 IU/mL containing 23        mM of m-cresol, 174 mM of glycerol and 260 μM of zinc chloride        at pH 7.4.

The clear solution is filtered (0.22 μm) and introduced into 3 mL glasscartridges for injector pen.

EXAMPLE CA4 Introduction into Cartridges of a Solution of pramlintide at0.6 mg/mL and of Human Insulin at 100 IU/mL in the Presence ofCo-Polyamino Acid AB24 at 2.4 mg/mL, m-cresol (29 mM), glycerol (174 mM)and zinc chloride (229 μM) at pH 6.6

The solution of pramlintide at 0.6 mg/mL and of human insulin at 100IU/mL in the presence of co-polyamino acid AB24 at 2.4 mg/mL, m-cresol(29 mM), glycerol (174 mM) and zinc chloride (229 μM) at pH 6.6described in example CF4 is filtered (0.22 μM) is introduced into 3 mLglass cartridges for injector pen.

EXAMPLE CA5 (Pump Stability) Preparation of a Solution of pramlintide at0.6 mg/mL and of Insulin Lispro at 100 IU/mL at pH 6.6 ContainingCo-Polyamino Acid AB24, m-cresol (29 mM), glycerol (174 mM) and zincchloride (300 μM)

A concentrated solution of co-polyamino acid AB24 and excipients isprepared by adding concentrated solutions of excipients (m-cresol,glycerol, zinc chloride) to a concentrated solution of co-polyamino acidAB24.

A concentrated solution of pramlintide at 10 mg/mL, then a solution ofinsulin lispro at 200 IU/mL are added to this concentrated solution ofco-polyamino acid AB24 and excipients in such a manner as to obtain theintended final composition. The final pH is adjusted to 6.6 by additionof NaOH/HCl.

EXAMPLE CA6 Preparation of a Solution of Human Insulin at 100 IU/mLContaining Co-Polyamino Acid BB15 (10 mg/mL), m-cresol (29 mM), glycerol(174 mM) and zinc chloride (229 μM) at pH 6.6

A concentrated solution of co-polyamino acid BB15 and excipients isprepared by adding concentrated solutions of excipients (m-cresol,glycerol, zinc chloride) to a concentrated solution of co-polyamino acidBB15.

A solution of human insulin at 800 IU/mL is added to this concentratedsolution of co-polyamino acid BB15 and excipients in such a manner as toobtain the intended final composition. The final pH is adjusted to 6.6by addition of NaOH/HCl. The clear solution is filtered (0.22 μM) andintroduced into 3 mL glass cartridges for injector pen.

EXAMPLE CA7 Preparation of a Solution of pramlintide at 0.6 mg/mL and ofHuman Insulin at 100 IU/mL Containing Co-Polyamino Acid BB15 (10 mg/mL),m-cresol (29 mM), glycerol (174 mM) and zinc chloride (229 μM) at pH 6.6

A concentrated solution of co-polyamino acid BB15 and excipients isprepared by adding concentrated solutions of excipients (m-cresol,glycerol, zinc chloride) to a concentrated solution of co-polyamino acidBB15.

A concentrated solution of pramlintide at 10 mg/mL, then a concentratedsolution of human insulin at 500 IU/mL are added to this concentratedsolution of co-polyamino acid BB15 and excipients in such a manner as toobtain the intended final composition. The final pH is adjusted to 6.6by addition of NaOH/HCl. The clear solution is filtered (0.22 μM) andintroduced into 3 mL glass cartridges.

D. Physicochemistry D I: Results of Visual Observations Made During theMixing and of Measurements of Fibrillation by ThT Principle

The poor stability of a peptide can lead to the formation of amyloidfibrils, which are defined as ordered macromolecular structures. Thesestructures may result in the formation of a gel within the sample.

The test of monitoring the fluorescence of Thioflavin T (ThT) is used toanalyze the physical stability of the solutions. Thioflavin is a smallprobe molecule having a characteristic fluorescence signature when itbinds to amyloid type fibrils (Naiki et al. (1989) Anal. BioChem. 177,244-249; LeVine (1999) Methods. Enzymol. 309, 274-284).

This method makes it possible to monitor the formation of fibrils forlow concentrations of ThT within undiluted solutions. This monitoring iscarried out under accelerated stability conditions: under stirring andat 37° C.

Experimental Conditions

The samples were prepared immediately before the start of themeasurement. The preparation of each composition is described in theassociated example. Thioflavin T is added to the composition from aconcentrated stock solution in such a manner as to induce a negligibledilution of the composition. The concentration of Thioflavin T in thecomposition is 1, 2 or 40 μM depending on the composition type: 40 μM inthe case of compositions of human amylin at 0.6 mg/mL, 2 μM in the caseof compositions of pramlintide at 0.9 mg/mL and 1 μM in the compositionsof pramlintide at 0.4 mg/mL. This concentration is indicated in thelegend pertaining to the table of results of the latency times for eachtype of composition.

A volume of 150 μL of the composition was introduced into a well of a96-well plate. Each composition was analyzed in three tests (triplicate)on the same plate. The plate was sealed with a transparent film in orderto prevent evaporation of the composition.

This plate was then placed into the enclosure of a plate reader(EnVision 2104 Multilabel, Perkin Elmer). The temperature is adjusted to37° C., and lateral agitation at 960 rpm with an amplitude of 1 mm isapplied.

A reading of the fluorescence intensity in each well is carried out atan excitation wavelength of 442 nm and an emission wavelength of 482 nmover time.

The fibrillation process manifests itself in a strong increase influorescence after a delay referred to as latency time.

For each well, this delay was determined graphically from theintersection between the baseline of the fluorescence signal and theslope of the fluorescence curve as a function of time determined duringthe strong initial increase in fluorescence. The value of the latencytime plotted corresponds to the average of the latency time measurementsperformed on three wells.

An example of a graphic determination is represented in FIG. 1.

This figure is a graphic representation of the determination of thelatency time (LT) by monitoring the fluorescence of Thioflavin T, on acurve having the fluorescence value (in u.a., arbitrary units) on theordinate and the time in minutes on the abscissa.

EXAMPLE D1 Stability of Solutions of Human Amylin at 0.6 mg/mL at pH 7.4in the Presence of Co-Polyamino Acid BB15 at Different Concentrations

TABLE 22 Measurement of the latency time by ThT (40 μM) of the solutionsCV1 to CV10. Ratio Concentration of BB15/human co-polyamino acid amylinBB15 Solution mol/mol mg/mL mM Latency time (h) CV1 — — — <0.02 CV5 5 30.73 >1 CV6 6 3.6 0.88 >5 CV7 7 4.2 1.03 >30 CV8 8 4.8 1.17 >54 CV9 95.4 1.32 >54 CV10 10 6 1.47 >72

The latency times of a solution of human amylin at pH 7.4 (CV1), withoutco-polyamino acid, is less than 0.02 h; the solutions CV5 to CV10according to the invention, containing molar ratios BB15/human amylingreater than 5 make it possible to obtain latency times of more than onehour, a molar ratio of 10 making it possible to obtain latency times ofmore than 72 h.

EXAMPLE D2 Stability of Solutions of Human Amylin at 0.6 mg/mL and ofHuman Insulin at 100 IU at pH 7.4 in the Presence of Co-Polyamino AcidBB15 at Different Concentrations

TABLE 23 Measurement of the latency time by ThT (40 μM) of thesolutions, CX6, CX10 and CX11. Ratio BB15/human Concentration of co-amylin polyamino acid BB15 Solution mol/mol mg/mL mM Latency time (h)CX1 — — — * CX6 6 3.6 0.88 >0.1 CX10 10 6 1.47 >0.5 CX11 17.5 10.52.57 >5 * Latency time not measured because of turbid solution.

The solution of human amylin and of human insulin at pH 7.4 (CX1) isturbid. The co-polyamino acid BB15 makes it possible to obtain a clearsolution containing human amylin in the presence of human insulin at pH7.4 with latency times of more than 0.1 hour starting with a molar ratioBB15/human amylin of 6, and latency times of more than 5 h for a molarratio of BB15/human amylin of 17.5.

EXAMPLE D3 Stability of Solutions of pramlintide at 0.4 mg/mL at pH 7.4in the Presence of Co-Polyamino Acid BB15 at Different Concentrations

TABLE 24 Measurement of the latency time by ThT (1 μM) of the solutionsCW1 to CW3. Concentration of co- Ratio polyamino acid BB15BB15/pramlintide Solution mg/mL mM mol/mol Latency time (h) CW1 — — 0.7CW2 2.4 0.59 6 >40 CW3 4 0.98 10 >63

The solution of pramlintide at pH 7.4 (CW1) without co-polyamino acidhas a short latency time. The co-polyamino acid BB15 makes it possibleto obtain a solution containing pramlintide at pH 7.4 with latency timesof more than 40 h starting with a molar ratio of BB15/pramlintide of 6.

EXAMPLE D4 Stability of Solutions of pramlintide at 0.9 mg/mL at pH 7.4in the Presence of Co-Polyamino Acid BB15 at Different Concentrations

TABLE 25 Measurement of the latency time by ThT (2 μM) of the solutionsCY1 to CY7 Ratio BB15/pramlintide Concentration of BB15 Solution mol/molmg/mL mM Latency time (h) CY1 — — — 0.7 CY2 2 1.8 0.44 >0.8 CY3 3 2.70.66 >4 CY4 4 3.6 0.88 >30 CY5 5 4.5 1.10 >63 CY6 6 5.4 1.22 >63 CY7 109 1.32 >63

The solution of pramlintide at pH 7.4 (CY1) without co-polyamino acidhas a short latency time; the latency times of solutions containing aco-polyamino acid are greater than or equal to the latency times of thecomposition without co-polyamino acid at a molar ratio of co-polyaminoacid BB15/pramlintide of 2:1.

EXAMPLE D5 Stability of Solutions of pramlintide at 0.9 mg/mL at pH 7.4in the Presence of Different Co-Polyamino Acids

TABLE 26 Measurement of the latency time by ThT (2 μM) of thecompositions CY1, CP2 to CP12. Ratio co-polyamino Concentration of co-acid/ Co- polyamino acid pramlintide Latency Solution polyaminoacidmg/mL mM mol/mol time (h) CY1 — — — <0.7 CP2 BB15 5 1.22 5.4 >63 10 2.4510.8 >63 CP3 BB14 10 1.96 8.6 >15 CP4 AB17 10 1.22 9.8 >63 CP10 BB18 50.72 3.2 >63 10 1.44 6.3 >63 CP11 BB9 5 1 4.4 >50 10 2.01 8.8 >63 CP12BB2 5 1.27 5.6 >10 10 2.54 11.2 >63

The solution of pramlintide at pH 7.4 (CY1) without co-polyamino acidhas a short latency time. The co-polyamino acids of the invention makeit possible to obtain latency times of more than 10 h under theconditions tested.

EXAMPLE D6 Stability of Solutions of pramlintide at 0.6 mg/mL at pH 6.6in the Presence of Co-Polyamino Acid BB15 at Different Concentrations

TABLE 27 Measurement of the latency time by ThT (2 μM) of the solutionCH1 and CH2 to CH8. Ratio Concentration of co- BB15/pramlintidepolyamino acid BB15 Latency time Solution mol/mol mg/mL mM (h) CH1 — — —1 CH2 2 1.3 0.29 >4 CH3 3 2 0.45 >10 CH4 4 2.7 0.61 >50 CH5 6 4 0.90 >50CH6 8 5.3 1.19 >50 CH7 10 6.7 1.50 >50 CH8 15 10 2.24 >50

The solution of pramlintide at pH 6.6 (CH1) without co-polyamino acidhas a short latency time; the latency times of the solutions containinga co-polyamino acid are greater than the latency times of thecomposition without co-polyamino acid at a molar ratio of co-polyaminoacid BB15/pramlintide of 2:1.

EXAMPLE D7 Stability of Solutions of pramlintide at 0.6 mg/mL at pH 6.6in the Presence of Different Co-Polyamino Acids

TABLE 28 Measurement of the latency time by ThT (2 μM) of thecompositions CI1 to CI14. Ratio Co- Concentration of co-polyaminopolyamino co-polyamino acid acid/pramlintide Latency Solution acid mg/mLmM mol/mol time (h) CI1 BB20 1.3 0.3 2 >10 2.6 0.6 4 >20 CI2 BB21 1.30.6 4 >10 CI3 AB22 2.4 0.3 2 >10 CI4 BB24 2.9 0.6 4 >10 CI5 BB25 1.5 0.32 >50 3 0.6 4 >50 CI6 AB23 3.4 0.23 2 >10 CI7 AB28 2.3 0.3 2 >10 4.7 0.64 >10 CI8 AB24 1.2 0.15 1 >10 2.4 0.3 2 >50 CI9 AB25 1.3 0.15 1 >10 2.60.3 2 >10 CI10 AB26 1.5 0.3 2 >10 CI11 AB27 1.3 0.15 1 >5 2.7 0.3 2 >10CI12 AB31 1.3 0.15 1 >10 2.5 0.3 2 >10 CI13 AB29 8.9 1.15 7.6 >5 CI14AB32 1.3 0.15 1 >10 2.5 0.3 2 >10

The solution of pramlintide at pH 6.6 (CH1) without co-polyamino acidhas a short latency time. The co-polyamino acids of the invention makeit possible to obtain a latency time of more than 5 h under theconditions tested.

EXAMPLE D7A Stability of Solutions of pramlintide at 0.6 mg/mL at pH 6.6in the Presence of Co-Polyamino acid AB14 and of Different Contents ofsodium chloride and of zinc chloride

TABLE 29 Measurement of the latency time by ThT (2 μM) of thecompositions BT1 to BT5 Concentration of Co- co-polyamino Latencypolyamino acid [NaCl] [ZnCl₂] time Solution acid mg/mL mM (mM) (mM) (h)CT1 AB14 6.3 1.87 — 0.75 0.6 CT2 AB14 6.3 1.87 50 — >2 CT3 AB14 6.3 1.87100 — >5 CT4 AB14 6.3 1.87 50 0.75 >5 CT5 AB14 6.3 1.87 100 0.75 >20

The solution of pramlintide at pH 6.6 and of co-polyamino acid AB14 hasa longer latency time in the presence of sodium chloride or of sodiumand zinc chloride.

EXAMPLE D7B Stability of Solutions of pramlintide at 0.6 mg/mL at pH 6.6in the Presence of Different Co-Polyamino Acids and of DifferentContents of sodium chloride and of zinc chloride

TABLE 30 Measurement of the latency time by ThT (2 μM) of thecompositions BS1 to BS11 Concentration of Latency Co-polyaminoco-polyamino acid [NaCl] [ZnCl₂] time Solution acid mg/mL mM (mM) (mM)(h) CS1 AB15 7.8 1.6 — — 3.5 CS2 AB15 11.7 2.3 — — >30 CS3 AB15 3.9 0.850 — >10 CS4 AB15 6.3 1.3 50 — >50 CS5 AB15 7.8 1.6 50 — >30 CS6 AB153.9 0.8 100 — >50 CS7 AB16 12.4 1.5 — — 8 CS8 AB16 16.7 2.1 — — >50 CS9AB16 7.4 0.9 50 — >20 CS10 AB16 12.4 1.5 50 — >50 CS11 AB16 7.4 0.9 50 1>30

The solutions of pramlintide at pH 6.6 and of co-polyamino acid AB15 andAB16 have a longer latency time in the presence of sodium chloride or ofsodium and zinc chloride.

EXAMPLE D8 Stability of Solutions of pramlintide at 0.4 mg/mL and ofHuman Insulin 100 IU/mL at pH 7.4 Containing Co-Polyamino Acid BB15

TABLE 31 Measurement of the latency time by ThT (1 μM) of thecompositions CN1 to CN3. Concentration of co- Ratio polyamino acid BB15BB15/pramlintide Solution mg/mL mM mol/mol Latency time (h) CN1 — — *CN2 2.4 0.59 6 >19 CN3 4 0.98 10 >19 * Latency time not measured becauseof turbid solution.

The solution of pramlintide and of human insulin at pH 7.4 (CN1) withoutcopolyamino acid is turbid.

Co-polyamino acid BB15 makes it possible to obtain a clear solution ofpramlintide at 0.4 mg/mL and of human insulin at 100 IU/mL at pH 7.4with latency times of more than 19 h for molar ratios BB15/pramlintidegreater than 6.

EXAMPLE D9 Stability of Solutions of pramlintide at 0.9 mg/mL and ofHuman Insulin 100 IU/mL at pH 7.4 in the Presence of Co-Polyamino AcidBB15 at Different Concentrations

TABLE 32 Measurement of the latency time by ThT (2 μM) of thecompositions CR1 to CR4 and CU3 to CU8. Concentration of Ratioco-polyamino acid BB15/pramlintide BB15 Solution mol/mol mg/mL mMLatency time (h) CR1 — — — * CU3 2 1.8 0.44 >0.5 CR2 3 2.7 0.66 >2 CR3 43.6 0.88 >6 CR4 5 4.5 1.10 >9 CU7 6 5.4 1.32 >9 CU8 10 9 2.20 >9 *Latency time not measured because of turbid solution.

A solution of pramlintide at 0.9 mg/mL and of human insulin 100 IU/mL atpH 7.4 (BR1) without co-polyamino acid is turbid. The clear solutions ofpramlintide at 0.9 mg/mL and of human insulin 100 IU/mL at pH 7.4 in thepresence of co-polyamino acid BB15 have latency times of more than 0.5hour at the molar ratio BB15/pramlintide of 2, which can be greater than9 h for molar ratios BB15/pramlintide greater than 5.

EXAMPLE D10 Stability of Solutions of pramlintide at 0.9 mg/mL and ofHuman Insulin 100 IU/mL at pH 7.4 in the Presence of DifferentCo-Polyamino Acids

TABLE 33 Measurement of the latency time by ThT (2 μM) of the solutionsCG2 to CG12. Ratio Co- Concentration of co- co-polyamino polyaminopolyamino acid acid/pramlintide Latency Solution acid mg/mL mM mol/moltime (h) CR1 — — * CG2 BB15 5 1.22 5.4 >9 10 2.45 10.8 >7 CG3 BB14 101.96 8.6 >9 CG4 AB17 5 1.11 4.9 >2 10 1.22 9.8 >5 CG5 AB15 10 1.998.8 >2 CG10 BB18 5 0.72 3.2 >1 10 1.44 6.3 >4 CG11 BB9 5 1 4.4 >4 102.01 8.8 >3 CG12 BB2 5 1.27 5.6 >5 10 2.54 11.2 >6 * Latency time notmeasured because of turbid solution.

The solution of pramlintide and of human insulin at pH 7.4 (CR1) isturbid. The co-polyamino acids make it possible to obtain latency timesof more than 1 hour under the conditions tested.

EXAMPLE D11 Stability of Solutions of pramlintide at 0.9 mg/mL and ofInsulin Lispro 100 IU/mL at pH 7.4 in the Presence of Co-Polyamino AcidBB15 at Different Concentrations

TABLE 34 Measurement of the latency time by ThT (2 μM) of the solutionsCD1 and CD3 to CD8. Concentration of co- Ratio polyaminoBB15/pramlintide acid BB15 Latency time Solution mol/mol mg/mL mM (h)CD1 — — * CD3 2 1.8 0.44 0.8 CD4 3 2.7 0.66 >2 CD5 4 3.6 0.88 >7 CD6 54.5 1.10 >9 CD7 6 5.4 1.22 >9 CD8 10 9 1.32 >9 *Latency time notmeasured because of turbid solution.

The solution of pramlintide and of insulin lispro at pH 7.4 (CD1) isturbid. The co-polyamino acids make it possible to obtain latency timesof more than 0.8 hour under the conditions tested.

EXAMPLE D12 Stability of Solutions of pramlintide at 0.6 mg/mL and ofHuman Insulin 100 IU/mL at pH 6.6 in the Presence of Co-Polyamino AcidBB15 at Different Concentrations

TABLE 35 Measurement of the latency time by ThT (2 μM) of the solutionsCK1 and CK3 to CK8. Concentration of co- Ratio polyaminoBB15/pramlintide acid BB15 Latency time Solution mol/mol mg/mL mM (h)CK1 — — — * CK3 3 2 0.45 >0.5 CK4 4 2.7 0.61 >5 CK5 6 4 0.90 >5 CK6 85.3 1.19 >5 CK7 10 6.7 1.50 >5 CK8 15 10 2.24 >5 *Latency time notmeasured because of turbid solution.

The solution of pramlintide and of human insulin at pH 6.6 (CK1) isturbid. The clear solutions of pramlintide at 0.6 mg/mL and of humaninsulin 100 IU/mL at pH 6.6 in the presence of co-polyamino acid BB15have latency times of more than 0.5 h at a molar ratio BB15/pramlintideof 3, which can be greater than 5 h starting with a ratioBB15/pramlintide of 4.

EXAMPLE D13 Stability of Solutions of pramlintide at 0.6 mg/mL and ofHuman Insulin 100 IU/mL at pH 6.6 in the Presence of DifferentCo-Polyamino Acids

TABLE 36 Measurement of the latency time by ThT (2 μM) of the solutionsCM1 to CM18. Concentration of co- Ratio polyamino co-polyamino LatencyCo-polyamino acid acid/pramlintide time Solution acid mg/mL mM mol/mol(h) CM1 BB20 2.6 0.61 4 >1 5.3 1.22 8 >1 CM2 BB21 1.3 0.6 4 >10 CM3 AB222.4 0.3 2 >5 CM4 BB24 2.9 0.6 4 >10 CM5 BB23 3 0.76 5 >10 CM6 BB25 1.50.3 2 >10 CM7 BB22 2.7 0.6 4 >5 CM8 AB23 7.7 0.69 4.6 >15 CM9 BB19 4.70.4 2.8 >1 CM10 AB28 2.3 0.3 2 >10 CM11 AB24 2.4 0.3 2 >5 CM12 AB25 2.60.3 2 >5 CM13 AB26 1.5 0.3 2 >1 2.3 0.5 3 >10 CM14 AB27 1.3 0.15 1 >12.7 0.3 2 >5 CM15 AB30 1.2 0.15 1 >1 2.3 0.3 2 >10 CM16 AB31 1.3 0.151 >1 2.5 0.3 2 >10 CM17 AB29 5.9 0.8 5 >1 8.9 1.15 7.6 >5 CM18 AB32 2.50.3 2 >1

The solution of pramlintide and of human insulin at pH 6.6 (CK1) isturbid. The co-polyamino acids make it possible to obtain latency timesof more than 1 h under the conditions tested.

EXAMPLE D13A Stability of Solutions of pramlintide at 0.6 mg/mL and ofHuman Insulin 100 IU/mL at pH 6.6 in the Presence of DifferentCo-Polyamino Acids and of Different Contents of sodium chloride and ofzinc chloride

TABLE 37 Measurement of the latency time by ThT (2 μM) of the solutionsBQ1 and BQ2 to BQ12 Co- Concentration of Latency polyamino co-polyaminoacid [NaCl] [ZnCl₂] time Solution acid mg/mL mM (mM) (mM) (h) BQ1 AB146.3 1.87 100 1 >5 BQ2 AB15 7.8 1.6 — 0.23 >2 BQ3 AB15 11.7 2.3 — 0.23 >2BQ5 AB15 6.3 1.3 50 0.23 >2 BQ6 AB15 7.8 1.6 50 0.23 >5 BQ7 AB15 3.9 0.8100 0.23 >2 BQ8 AB15 6.3 1.3 100 0.23 >2 BQ9 AB15 7.8 1.6 100 0.23 >5BQ10 AB16 7.4 0.9 50 0.23 >1 BQ11 AB16 12.4 1.5 50 0.23 >2 BQ12 AB16 7.40.9 50 1 >2

The solutions of pramlintide and of human insulin at pH 6.6 in thepresence of the co-polyamino acids AB14, AB15 and AB16, of sodium andzinc chloride have latency times of more than 1 h under the conditionedtested. The addition of sodium chloride or sodium and zinc chloridemakes it possible to increase the latency times.

EXAMPLE D14 Stability of Compositions Having Variable pramlintideConcentrations and Human Insulin at 100 IU/mL in the Presence ofCo-Polyamino Acid AB24, m-cresol (29 mM), glycerol (174 mM) and zincchloride (229 μM) at pH 6.6

TABLE 38 Measurement of the latency time by ThT (2 μM) of solutions CF2to CF6. Concentration of co- Ratio Concentration polyamino acidco-polyamino Latency of pramlintide AB24 acid/pramlintide time Solution(mg/mL) mg/mL mM mol/mol (h) CF2 0.9 5.4 0.67 3 >10 (12)   CF3 0.8 4.80.6 3 >10 (14.1) CF4 0.6 3.6 0.45 3 >5 (5.5) CF5 0.3 1.8 0.22 3 >5 (5.6)CF6 0.2 1 0.125 2.5 >5 (5.4)

The solutions of pramlintide at variable concentrations and of humaninsulin at 100 IU/mL at pH 6.6 are turbid (examples CF1A-E). Thesolutions of pramlintide at variable concentrations and of human insulinat 100 IU/mL at pH 6.6 in the presence of co-polyamino acid AB24 havelatency times of more than 5 h under the conditions tested.

D II: Study of the Stability of the Compositions According to theInvention D II A: Preparation of the Compositions

Composition D1: Preparation of a Solution of pramlintide at 0.9 mg/mLContaining m-cresol (29 mM) and glycerol (174 mM) at pH 6.6.

By a method similar to the one used in example CH1, a solution ofpramlintide at 0.9 mg/mL containing m-cresol (29 mM) and glycerol (174mM) at pH 6.6 is obtained. The solution is clear.

Composition D2: Preparation of a Solution of pramlintide at 0.9 mg/mLContaining Co-Polyamino Acid BB15, m-cresol (29 mM) and glycerol (174mM) at pH 6.6.

By a method similar to the one used in example BH0, a solution ofpramlintide at 0.9 mg/mL and of co-polyamino acid BB15 at 10 mg/mLcontaining m-cresol (29 mM) and glycerol (174 mM) at pH 6.6 is obtained.The solution is clear.

D II B: Procedure for Visual Inspection:

The 3 mL vials or cartridges filled with 1 mL of formulation areinspected visually in order to detect the appearance of visibleparticles or of turbidity. This inspection is carried out according tothe recommendations of the European pharmacopoeia (EP 2.9.20): the vialsare subjected to elimination of at least 2000 lux and are observed on awhite background and a black background. The number of weeks or monthsof stability corresponds to the duration after which the solutionscontain visible particles or are turbid.

These results are in agreement with the US pharmacopoeia (USP <790>).

D II C: Procedure for Assaying the Formulations:

The quantification of the purity of pramlintide and of insulin and ofthe recovery of native peptide is carried out by reversed-phase HPLC,provided with a CA18 column having the dimensions 4.6×150 mm with aparticle size of 3.5 μm. The pramlintide is detected at a wavelength of214 nm, and the insulin is detected at a wavelength of 276 nm. Theelution is carried out in an aqueous mobile phase with a linearacetonitrile gradient.

The recovery of pramlintide or of insulin (%) at time t represents theratio between the area under the peak of pramlintide or the area underthe peak of insulin at time t and the area of the initial pramlintidepeak.

The purity of pramlintide and of insulin (%) represents the ratiobetween the area of the absorbance peak of pramlintide or of insulin andthe total area of all the peaks including pramlintide and itsimpurities.

D II D: Physical Stability in Cartridges at 37° C. of Solutions ofpramlintide at 0.9 mg/mL in the Presence of Co-Polyamino acid BB15,m-cresol (29 mM) and glycerol (174 mM) at pH 6.6 or pH 7.4.

The solutions D1, CY1, D2 and CY7 are filtered (0.22 μm). 1 mL ofsolution is introduced into a 3 mL glass cartridge for auto-injectorpen. The cartridges are placed in an oven at 37° C. under staticconditions. The cartridges are observed at a weekly frequency.

TABLE 39 Results of the physical stabilities at 37° C. in cartridges ofthe compositions of pramlintide at 0.9 mg/mL in the presence ofco-polyamino acid BB15 Physical Concentration of co-polyamino stabilityacid BB15 37° C. in cartridges Solution mg/mL pH (weeks) D1 — 6.6 <1 CY1— 7.4 <1 D2 10 6.6 >4 CY7 10 7.4 >4

The solutions of pramlintide at 0.9 mg/mL at pH 6.6 and pH 7.4 have aphysical stability at 37° C. in a cartridge of less than one week.

The solutions of pramlintide at 0.9 mg/mL at pH 6.6 and pH 7.4 in thepresence of co-polyamino acid BB15 have a physical stability at 37° C.in cartridges of at least 4 weeks.

EXAMPLE D II C Chemical Stability in Cartridges at 37° C. of Solutionsof pramlintide at 0.9 mg/mL in the Presence of Co-Polyamino Acid BB15,m-cresol (29 mM) and glycerol (174 mM) at pH 6.6 and 7.4

The solutions described in example D II D are analyzed by RP-HPLCchromatography.

TABLE 40 Results of the chemical stabilities of the compositions ofpramlintide at 0.9 mg/mL in the presence of co-polyamino acid BB15.Concentration of Purity co-polyamino Recovery pramlintide (%) acid BB15pramlinitide (%) 32 days Solution mg/mL pH 32 days - 37° C. T0 37° C. D1— 6.6 <60 97.2 <50 CY1 — 7.4 <20 94.7 <50 D2 10 6.6 >90 97.8 >60 CY7 107.4 >60 98.6 >60

The solutions of pramlintide at 0.9 mg/mL at pH 6.6 and pH 7.4 presenthave a recovery of pramlintide less than 60% % and the purity ofpramlintide is less than 50% % after 32 days at 37° C. in a cartridge.

The solutions of pramlintide at 0.9 mg/mL at pH 6.6 and pH 7.4 in thepresence of co-polyamino acid BB15 have a recovery of more than 65% %and can be greater than 90% % at pH 6.6 after 32 days at 37° C. incartridges. In the presence of co-polyamino acid BB15, the purity ofpramlintide is greater than 65% and can be greater than 85% at pH 6.6.

EXAMPLE D II E Physical Stability in a Vial and Cartridge at 30° C. ofSolutions of pramlintide at 0.9 mg/mL and at 0.6 mg/mL in the Presenceof Co-Polyamino Acid BB15, m-cresol (29 mM) and glycerol (174 mM) at pH6.6

The solutions D1, CH1, D2 and CH8 are filtered (0.22 μm). 1 mL ofsolution is introduced into 3 mL glass cartridges for auto-injector penand in 3 mL glass vials. The cartridges and the vials are placed in anoven at 30° C. under static conditions, then observed every 2 weeks.

TABLE 41 Results of the physical stabilities in a vial and in acartridge at 30° C. of the compositions of pramlintide at 0.9 and 0.6mg/mL in the presence of co-polyamino acid BB15 Physical PhysicalConcentration stability stability co-polyamino Concentration 30° C. 30°C. BB15 acid Pramlintide in vial in cartridge Solution mg/mL (mg/mL) pH(weeks) (weeks) D1 — 0.9 6.6 <7 <2 CH1 — 0.6 6.6 <7 — D2 10 0.96.6 >12 >12 CH8 10 0.6 6.6 >12 >12

The solutions of pramlintide at 0.9 mg/mL and 0.6 mg/mL at pH 6.6 have aphysical stability in a vial of less than 7 weeks at 30° C. The physicalstability in a cartridge of the solution of pramlintide at 0.9 mg/mL pH6.6 is less than 2 weeks.

The solutions of pramlintide at 0.9 mg/mL and 0.6 mg/mL at pH 6.6 in thepresence of co-polyamino acid BB15 have a physical stability at 30° C.of more than 12 weeks in a vial and in a cartridge.

EXAMPLE D II F Chemical Stability in a Vial at 30° C. of Solutions ofpramlintide at 0.9 mg/mL and at 0.6 mg/mL in the Presence ofCo-Polyamino Acid BB15, m-cresol (29 mM) and glycerol (174 mM) at pH 6.6

The solutions described in example D II E are analyzed by RP-HPLCchromatography.

TABLE 42 Results of the chemical stabilities in a vial at 30° C. of thecompositions of pramlintide at 0.9 and 0.6 mg/mL in the presence ofco-polyamino acid BB15 at pH 6.6. Recovery Concentration pramlintidePurity co-polyamino Concentration (%) pramlintide (%) acid BB15pramlintide 5 weeks 5 weeks Solution mg/mL (mg/mL) 30° C. T0 30° C. D1 —0.9 <70 97.2 <60 D2 10 0.9 >95 97.8 >90 CH8 10 0.6 >95 94.6 >90

The solution of pramlintide at 0.9 mg/mL at pH 6.6 has a recovery ofpramlintide of less than 70% and the purity of pramlintide is less than60% after 5 weeks at 30° C. in a vial.

The solutions of pramlintide at 0.9 mg/mL and 0.6 mg/mL at pH 6.6 in thepresence of co-polyamino acid BB15 have a recovery of pramlintidegreater than 95% and the purity of pramlintide is greater than 90% after5 weeks at 30° C.

EXAMPLE D II F Physical Stability in a Vial and Cartridges at 30° C. ofSolutions of pramlintide at 0.6 mg/mL and of Insulin 100 IU/mL at pH 6.6in the Presence of Co-Polyamino Acid BB15, m-cresol (29 mM), glycerol(174 mM) and zinc at pH 6.6

The solution CK8 is filtered (0.22 μM). 1 mL of solution is introducedinto 3 mL glass cartridges for auto-injector pen and in 3 mL glassvials. The cartridges and the vials are placed in an oven at 30° C.under static conditions, then observed every 2 weeks.

TABLE 43 Results of the physical stabilities in a vial and in acartridge at 30° C. of the compositions of pramlintide at 0.6 mg/mL, ofinsulin 100 IU/mL, and in the presence of co-polyamino acid BB15 at pH6.6. Physical Physical Concentration stability stability co-polyaminoConcentration Concentration 30° C. 30° C. in acid BB15 pramlintideInsulin in vial cartridge Solution mg/mL (mg/mL) (IU/mL) (weeks) (weeks)CK1 — 0.6 100 * * CK8 10 0.6 100 >3 >12 *solution turbid from itspreparation on.

The solution of pramlintide at 0.6 mg/mL and of insulin at 100 IU/mL atpH 6.6 is turbid.

The solution of pramlintide at 0.6 mg/mL and of human insulin at 100IU/mL at pH 6.6 in the presence of co-polyamino acid BB15 has a physicalstability at 30° C. of more than 3 weeks in a vial and more than 12weeks in a cartridge.

EXAMPLE D II G Chemical Stability in a Vial at 30° C. of a Solution ofpramlintide at 0.6 mg/mL and of Insulin 100 IU/mL at pH 6.6 in thePresence of Co-Polyamino Acid BB15, m-cresol (29 mM), glycerol (174 mM)and zinc

The solution described in example D II F is analyzed by RP-HPLCchromatography.

TABLE 44 Results of chemical stability in a vial at 30 ?+0C of acomposition of pramlintide at 0.6 mg/mL, of insulin 100 IU in thepresence of co-polyamino acid BB15 at pH 6.6. Recovery Purity RecoveryPurity pramlintide pramlintide (%) Insulin (%) Insulin (%) (%) 5 weeks 5weeks 5 weeks Solution 5 weeks 30° C. T0 30° C. 30° C. T0 30° C. CK8 >9096.8 >90 >90 97.9 >90

The solution of pramlintide at 0.6 mg/mL and of insulin at 100 IU/mL atpH 6.6 in the presence of co-polyamino acid BB15 has a recovery ofpramlintide of more than 90% and the purity of pramlintide is greaterthan 90% after 5 weeks at 30° C. in a vial. The recovery of insulin isgreater than 90% and the purity of the insulin is greater than 90% after5 weeks at 30° C. in a vial.

EXAMPLE D II H Physical Stability in a Vial at 30° C. and in a Cartridgeat 30° C137° C. of Solutions of pramlintide at 0.6 mg/mL and of Insulin100 IU/mL at pH 6.6 in the Presence of Co-Polyamino Acid AB24 at 2.4mg/mL, m-cresol (29 mM), glycerol (174 mM) and zinc at pH 6.6

Solution CM11 is filtered (0.22 μM). 1 mL of solution is introduced into3 mL glass cartridges for auto-injector pen and into 3 mL glass vials.The cartridges and the vials are placed in an oven at 30° C. understatic conditions, then observed every 2 weeks. Cartridges are alsoplaced in an oven at 37° C. under static conditions, then observed everyweek.

TABLE 45 Results of the physical stabilities in a vial at 30° C. and ina cartridge at 30 and 37° C. of the compositions of pramlintide at 0.6mg/mL, of insulin 100 IU/mL and in the presence of co-polyamino acidAB24 at pH 6.6. Physical Physical Physical Concentration stabilitystability stability co-polyamino Concentration Concentration 30° C. 30°C. in 37° C. in acid AB24 pramlintide Insulin in vial cartridgecartridge Solution mg/mL (mg/mL) (IU/mL) (weeks) (weeks) (weeks) CK1 —0.6 100 * * * CM11 2.4 0.6 100 >9 >12 >9 * solution turbid from itspreparation on.

The solution of pramlintide at 0.6 mg/mL and of insulin at 100 IU/mL atpH 6.6 is turbid.

The solution of pramlintide at 0.6 mg/mL and of human insulin at 100IU/mL at pH 6.6 in the presence of co-polyamino acid AB24 has a physicalstability at 37° C. of more than 9 weeks in a vial and of more than 12weeks in a cartridge. The physical stability at 37° C. in a cartridge isgreater than 9 weeks.

EXAMPLE D II I Chemical Stability in a Vial and Cartridge at 30° C. ofSolutions of pramlintide at 0.6 mg/mL and of Insulin 100 IU/mL at pH 6.6in the Presence of Co-Polyamino Acid AB24 at 2.4 mg/mL, m-cresol (29mM), glycerol (174 mM) and zinc at pH 6.6

The solution described in example D II H is analyzed by RP-HPLCchromatography.

TABLE 46 Results of the chemical stabilities in a vial and cartridge at30° C. of the compositions of pramlintide at 0.6 mg/mL, of insulin 100IU in the presence of co-polyamino acid AB24 at pH 6.6. Purity RecoveryPurity Recovery Insulin pramlintide pramlintide Insulin (%) (%) (%) (%)9 Solution 9 weeks 9 weeks 9 weeks weeks CM11 30° C. T0 30° C. 30° C. T030° C. Vial >88 96.7 >90 >90 97.4 >90 Cartridge >88 96.9 >85 >90 97.7>90

The solution of pramlintide at 0.6 mg/mL and of human insulin at 100IU/mL at pH 6.6 in the presence of co-polyamino acid AB24 has a recoveryof pramlintide greater than 88% and a purity greater than 90 and 85%,respectively, after 9 weeks of storage at 30° C. in a vial and in acartridge. Under these conditions, the recovery of insulin is greaterthan 90% and the purity of the insulin is greater than 90% in a vial andin a cartridge.

EXAMPLE D II J Physical Stability in a Cartridge at 4° C. of solutionsof pramlintide at 0.6 mg/mL at pH 6.6 in the Presence of Co-PolyaminoAcid BB15, m-cresol (29 mM), glycerol (174 mM) at pH 6.6

Solution CH8 is filtered (0.22 μm). 1 mL of solution is introduced into3 mL glass cartridges for auto-injector pen. The cartridges are placedin a refrigerator at 4° C.

TABLE 47 Results of the physical stability in a cartridge at 4° C. of acomposition of pramlintide at 0.6 mg/mL in the presence of co-polyaminoacid BB15 at pH 6.6. Physical stability Concentration co- Concentration4° C. polyamino acid BB15 pramlintide in cartridge Solution mg/mL(mg/mL) pH (month) CH8 10 0.6 6.6 >6

The solution of pramlintide at 0.6 mg/mL and of human insulin at 100IU/mL at pH 6.6 in the presence of co-polyamino acid BB15 has a physicalstability in a cartridge of more than 6 months.

EXAMPLE D II K Physical Stability in a Cartridge at 4° C. of Solutionsof pramlintide at 0.6 mg/mL at pH 6.6 and pH 7.4 in the Presence of DMPGat 4.5 mM

The solutions of CZ0 and CZ1 are filtered (0.22 μM). 1 mL of solution isintroduced into 3 mL glass cartridges for auto-injector pen. Thecartridges are placed in a refrigerator at 4° C.

TABLE 48 Results of the physical stabilities in a cartridge at 4° C. ofthe compositions of pramlintide at 0.6 mg/mL and in the presence of DMPGat pH 6.6 and 7.4. Physical stability at Concentration 4° C. in DMPGpramlintide cartridges Solution (mM) (mg/mL) pH Excipients (month) CZ04.5 0.6 6.6 m-cresol 29 mM <1.5 Glycerol 174 mM CZ1 4.5 0.6 7.4 phenol30 mM <1.5 Glycylglycine 8 mM pH 7.4 Glycerol 174 mM

The solutions of pramlintide at 0.6 mg/mL at pH 6.6 and at pH 7.4 have aphysical stability at 4° C. and in cartridges of less than 1.5 month(turbid solutions).

EXAMPLE D II L Pump Stability of Solutions of pramlintide at 0.6 mg/mLand of Human Insulin at 100 IU/mL at pH 6.6 in the Presence ofCo-Polyamino Acid AB24 at 3.6 mg/mL

The solution CM11 consisting of 0.6 mg/mL of pramlintide and 100 IU/mLof human insulin at pH 6.6 in the presence of co-polyamino acid AB24 at3.6 mg/mL is filtered (0.22 μm) and introduced into a 3 mL reservoir forinsulin pump (Minimed 530G system manufactured by Medtronic). The pumpis provided with an infusion set (Quick set Paradigm 9/100 manufacturedby Medtronic).

The insulin pump is placed in an oven at 37° C. on an orbital stirreradjusted at a speed of 100 rpm. The pump is adjusted to a basal flowrate of 0.8 IU/h. Bolus injections of 6 IU are performed 3 times per dayfor a total duration of 8 days.

Table 49 presents the results of the measurements of MFI (Micro-FlowImaging) and the assays carried out by RP-HPLC on the fractionscollected between the 7^(th) day and the 8^(th) day of stabilitytesting.

TABLE 49 Results of the chemical pump stability at 37° C. of thecompositions of pramlintide at 0.6 mg/mL, of insulin 100 IU/mL and inthe presence of co-polyamino acid AB24 at pH 6.6. Recovery Subvisibleparticles Recovery Purity Human Purity (Micro Flow pramlintidepramlintide insulin Insulin Imaging. MFI) (%) (%) (%) (%) 1 week 1 week1 week 1 week 1 week Solution 37° C. 37° C. T0 37° C. 37° C. T0 37° C.CM11 particles > 10 μM >95 97.3 >95 >95 97.4 >95 <6000 particles percontainer* particles > 25 μM  <600 particles per container* *StandardUSP <788> on the number of subvisible particles in the products forparenteral injection.

After one week of pump stability at 37° C., the solution of pramlintideat 0.6 mg/mL and of human insulin at 100 IU/mL at pH 6.6 in the presenceof co-polyamino acid AB24 is clear and has a number of subvisibleparticles in compliance with the standard USP <788>. Under theseconditions, the recovery and the purities of pramlintide and of insulinare greater than 95%.

EXAMPLE D II M Pump Stability of Solutions of pramlintide at 0.6 mg/mLand of Insulin Lispro at 100 IU/mL at pH 6.6 in the Presence ofCo-Polyamino Acid AB24 at 3.6 mg/mL

The solution CA5 consisting of 0.6 mg/mL of pramlintide and of 100 IU/mLof insulin lispro at pH 6.6 in the presence of co-polyamino acid AB24 at3.6 mg/mL is filtered (0.22 μm) and subjected to a pump stability testusing a protocol identical to the one described in example DC11.

TABLE 50 Results of the chemical pump stability at 37° C. of thecompositions of pramlintide at 0.6 mg/mL, of insulin lispro at 100 IU/mLand in the presence of co-polyamino acid AB24 at pH 6.6. Subvisibleparticles Recovery Purity Recovery Purity (Micro Flow pramlintidepramlintide lispro Insulin Imaging. MFI) (%) (%) (%) (%) 1 week 1 week 1week 1 week 1 week Solution 37° C. 37° C. T0 37° C. 37° C. T0 37° C. CA5particles > 10 μM >95 97.2 >95 >95 99.3 >95 <6000 particles percontainer* particles > 25 μM  <600 particles per container* *USP <788>criterion for the number of subvisible particles in the products forparenteral injections.

After one week of pump stability at 37° C., the solution of pramlintideat 0.6 mg/mL and of insulin lispro at 100 IU/mL at pH 6.6 in thepresence of co-polyamino acid AB24 is clear and has a number ofsubvisible particles in compliance with the USP <788> standard. Underthese conditions of recovery and the purities of pramlintide and insulinlispro are greater than 95%.

E. Pharmacodynamics and Pharmacokinetics

E1: Protocol of the Measurement of the Pharmacokinetics of Formulationsof pramlintide and of Insulin.

Domestic pigs weighing approximately 50 kg, which had been catheritizedbeforehand at the jugular, are fasted for 2.5 hours before the start ofthe experiment. In the hour preceding the injection of the formulationsof pramlintide and of insulin, 3 blood samples were collected in orderto determine the basal level of pramlintide.

The injection of formulations at the dose of 1.125 μg/kg for pramlintideand 0.1875 IU/kg for insulin is carried out subcutaneously in the flankof the animal using an insulin pen (Novo, Sanofi or Lilly) equipped witha 31 G needle.

Blood samples are then collected every 4 minutes for 20 minutes, thenevery 10 to 60 minutes up to 3 hours. After each sample collection, thecatheter is rinsed with a diluted heparin solution.

The blood so drawn is collected in a K2EDTA tube and centrifuged toisolate the plasma. The pramlintide levels in the plasma samples aremeasured using the sandwich ELISA immune-enzymatic method for eachanimal.

The pharmacokinetics curves expressed in delta of the basal level arethen traced.

The following pharmacokinetics parameters are then determined bynon-compartmental analysis using the software Phoenix WinNonlin:

-   -   t_(max) pramlintide corresponding to the time needed to reach        the maximum concentration of pramlintide in the plasma;    -   AUC_(Pram 0-30 min) corresponding to the area under the curve of        the pramlintide concentrations as a function of time between 0        and 30 minutes post-administration;    -   AUC_(Pram 60-180 min) corresponding to the area under the curve        of the concentrations of pramlintide as a function of time        between 60 and 180 minutes post-administration;    -   C_(last) corresponding to the last quantifiable pramlintide        concentration in the plasma;    -   t_(last) corresponding to the time at which C_(last) is        observed.

t_(max) is routinely used for evaluating the start of the absorption.AUC_(Pram 0-30 min) is routinely used for evaluating the early exposureto pramlintide in the plasma. As to AUC_(Pram 60-180 min), it makes itpossible to evaluate the late exposure to pramlintide in the plasma.C_(last) and t_(last) make it possible to study the late concentrationlevels.

E2: Pharmacokinetic Results of Pramlintide of the Formulations ofpramlintide and of Insulin of Examples CA2 and CA3

Pramlintide Number of Example Rh-Insulin co-polyamino acid (mg/mL) pigsCA1/CA2 100 — 1 8 (double CA3 100 BB15 0.6 10

The pharmacokinetic results of pramlintide, which are obtained with thecompositions described in examples CA1/CA2 and CA3, are presented inFIG. 2. The analysis of these profiles indicates that the composition ofexample CA3 comprising co-polyamino acid BB15, 100 IU/mL of insulin and0.6 mg/mL of pramlintide (curve traced with the squares corresponding toexample CA3) makes it possible to obtain an absorption of pramlintidewhich is slower than the absorption of the composition of the examplewith double injection comprising only pramlintide and insulin (curvetraced with the triangles corresponding to the double-injection examplesCA1/CA2). The pharmacokinetics parameters of pramlintide are given inthe following table:

t_(max) C_(last) t_(last) pramlintide AUC_(Pram 0-30 min)AUC_(Pram 60-180 min) pramlintide pramlintide Example (min) (min *pmol/L) (min * pmol/L) (min) (min) CA1/CA2 20 ± 10 2076 ± 1596 3286 ±1951 12 ± 5  153 ± 40 CA3 77 ± 44 1006 ± 885  5989 ± 3146 28 ± 24 168 ±25E3: Pharmacokinetic Results of the pramlintide of the Formulations ofpramlintide and of Insulin of Examples CA1/CA2 and CA4.

Rh-Insulin Pramlintide Number of Example (IU/mL) co-polyamino acid(μg/mL) pigs CA1/CA2 100 — — 8 CA4 100 AB24 0.6 12

The pharmacokinetic results of pramlintide obtained with thecompositions described in examples CA1/CA2 and CA4 are presented in FIG.3. The analysis of these profiles indicates that the composition ofexample CA4 comprising co-polyamino acid AB24, 100 IU/mL of insulin and0.6 μg/mL of pramlintide (curve traced with the squares corresponding toexample CA4) makes it possible to obtain an absorption of pramlintidewhich is slower than the absorption of the composition of the examplewith double injection comprising only pramlintide and insulin (curvetraced with the triangles corresponding to the double-injection examplesCA1/CA2). The pharmacokinetic parameters of pramlintide are reported inthe following table:

t_(max) C_(last) t_(last) pramlintide AUC_(Pram 0-30 min)AUC_(Pram 60-180 min) pramlintide pramlintide Example (min) (min *pmol/L) (min * pmol/L) (min) (min) CA1/CA2 20 ± 10 2076 ± 1596 3286 ±1951 12 ± 5  153 ± 40 CA4 68 ± 23 749 ± 316 8064 ± 2963 34 ± 22 175 ± 17

1. A co-polyamino acid bearing carboxylate charges and hydrophobic radicals Hy, said co-polyamino acid consisting of glutamic or aspartic units and said hydrophobic radicals Hy having the following formula I:

in which GpR is a radical of formula II or II′:

GpA is a radical of formula III or III′:

GpC is a radical of formula IV:

the * indicate the sites of attachment of the different groups; a is a whole number equal to 0 or to 1; b is a whole number equal to 0 or to 1; p is a whole number equal to 1 or 2 and if p is equal to 1 then a is equal to 0 or to 1 and GpA is a radical of formula III′, and if p is equal to 2 then a is equal to 1, and GpA is a radical of formula III; c is a whole number equal to 0 or to 1, and if c is equal to 0 then d is equal to 1 or to 2; d is a whole number equal to 0, to 1 or to 2; r is a whole number equal to 0 or to 1, and if r is equal to 0 then the hydrophobic radical of formula I is attached to the co-polyamino acid via a covalent bond between a carbonyl of the hydrophobic radical and a nitrogen atom in N-terminal position of the co-polyamino acid, thus forming an amide function originating from the reaction of an amine function in N-terminal position of the precursor of the co-polyamino acid and an acid function borne by the precursor of the hydrophobic radical, and if r is equal to 1 then the hydrophobic radical of formula I is attached to the co-polyamino acid: via a covalent bond between a nitrogen atom of the hydrophobic radial and a carbonyl of the copolyamino acid, thus forming an amide function originating from the reaction of an amine function of the precursor of the hydrophobic radical and an acid function borne by the precursor of the co-polyamino acid, or via a covalent bond between a carbonyl of the hydrophobic radical and a nitrogen atom in N-terminal position of the co-polyamino acid, thus forming an amide function originating from the reaction of an acid function of the precursor of the hydrophobic radical and an amine function in N-terminal position borne by the precursor of the co-polyamino acid; R is a radical selected from the group consisting of: a linear or branched divalent alkyl radical comprising 2 to 12 carbon atoms if GpR is a radical of formula II or 1 to 11 carbon atoms if GpR is a radical of formula II′ or II″; a linear or branched divalent alkyl radical comprising 2 to 11 carbon atoms if GpR is a radical of formula II or 1 to 11 carbon atoms if GpR is a radical of formula II′ or II″, 1 to 11 carbon atoms, said alkyl radical bearing one or more —CONH2 functions, and an unsubstituted ether or polyether radical comprising 4 to 14 carbon atoms and 1 to 5 oxygen atoms; A is a linear or branched alkyl radical comprising 1 to 6 carbon atoms; B is a linear or branched alkyl radical, optionally comprising an aromatic ring, comprising 1 to 9 carbon atoms; Cx is a linear or branched monovalent alkyl radical, in which x indicates the number of carbon atoms and: if p is equal to 1, x is from 11 to 25 (11≤x≤25): if p is equal to 2, x is from 9 to 15(9≤x≤15), the ratio i between the number of hydrophobic radicals and the number of glutamic or aspartic units being from 0 to 0.5 (0<i≤0.5); when several hydrophobic radicals are borne by a co-polyamino acid, then they are identical or different, the degree of polymerization DP of glutamic or aspartic units is from 5 to 250; the free acid functions being in the form of a salt of an alkali cation selected from the group consisting of Na+ and K+.
 2. The co-polyamino acid according to claim 1, wherein said hydrophobic radicals are selected from the hydrophobic radicals of formula I in which p=1, represented by the following formula V:

in which GpR, GpA, GpC, r and a have the definitions given above.
 3. The co-polyamino acid according to claim 1, wherein said hydrophobic radicals are selected from the hydrophobic radicals of formula I in which a=1 and p=2, represented by the following formula VI:

in which GpR, GpA, GpC and r have the definitions given above.
 4. The co-polyamino acid according to claim 1, wherein said co-polyamino acid bearing carboxylate charges and hydrophobic radicals is selected from the co-polyamino acids having the following formula VII:

in which, D represents, independently, either a —CH₂— group (aspartic unit) or a —CH₂—CH₂— group (glutamic unit), Hy is a hydrophobic radical selected from the hydrophobic radicals of formula I, V or VI in which r=1 and GpR is a radical of formula II, R₁ is a hydrophobic radical selected from the hydrophobic radicals of formula I, V or VI in which r=0 or r=1 and GpR is a radical of formula II′, or a radical selected from the group consisting of an H, a linear C2 to C10 acyl group, a branched C3 to C10 acyl group, benzyl, a terminal “amino acid” unit, and a pyroglutamate, R₂ is a hydrophobic radical selected from the hydrophobic radicals of formula I, V or VI in which r=1 and GpR is a radical of formula II, or a —NR′R″ radical, R′ and R″, which may be identical or different, being selected from the group consisting of H, the linear or branched or cyclic C2 to C10 alkyls, benzyl, and said alkyls R′ and R″ optionally forming together one or more saturated, unsaturated and/or aromatic carbon rings and/or optionally comprising heteroatoms selected from the group consisting of O, N and S, X represents an H or a cationic entity selected from the group comprising the metal cations; n+m represents the degree of polymerization DP of the co-polyamino acid, that is to say the average number of monomeric units per co-polyamino acid chain, and 5 n+m≤250.
 5. The co-polyamino acid according to claim 4, wherein said co-polyamino acid bearing carboxylate charges and hydrophobic radicals is selected from the co-polyamino acids of formula VII in which R₁═R′₁ and R₂═R′₂, having the following formula VIIa:

in which, m, n, X, D and Hy have the definitions given above; R′₁ is a radical selected from the group consisting of an H, a linear C2 to C10 acyl group, a branched C3 to C10 acyl group, benzyl, a terminal “amino acid” unit, and a pyroglutamate; R′₂ is a —NR′R″ radical, R′ and R″, which may be identical or different, being selected from the group consisting of H, the linear or branched or cyclic C2 to C10 alkyls, benzyl, and said alkyls R′ and R″ optionally forming together one or more saturated, unsaturated and/or aromatic carbon rings and/or optionally comprising heteroatoms selected from the group consisting of O, N and S.
 6. The co-polyamino acid according to claim 4, wherein said co-polyamino acid bearing carboxylate charges and hydrophobic radicals is selected from the co-polyamino acids of formula VII in which n=0, having the following formula VIIb:

in which m, X, D, R₁ and R₂ have the definitions given above and at least R₁ or R₂ is a hydrophobic radical of formula I, V or VI.
 7. A Hydrophobic radical Hy having the following formula I:

in which GpR is a radical of formula II or II′:

GpA is a radical of formula III or III′:

GpC is a radical of formula IV:

the * indicate the sites of attachment of the different groups; a is a whole number equal to 0 or to 1; b is a whole number equal to 0 or to 1; p is a whole number equal to 1 or 2 and if p is equal to 1 then a is equal to 0 or to 1 and GpA is a radical of formula III′, and if p is equal to 2 then a is equal to 1, and GpA is a radical of formula III; c is a whole number equal to 0 or to 1, and if c is equal to 0 then d is equal to 1 or to 2; d is a whole number equal to 0, to 1 or to 2; r is a whole number equal to 0 or to 1, and if r is equal to 0 then the hydrophobic radical of formula I is attached to the co-polyamino acid via a covalent bond between a carbonyl of the hydrophobic radical and a nitrogen atom in N-terminal position of the co-polyamino acid, thus forming an amide function originating from the reaction of an amine function in N-terminal position of the precursor of the co-polyamino acid and an acid function borne by the precursor of the hydrophobic radical, and if r is equal to 1 then the hydrophobic radical of formula I is attached to the co-polyamino acid: via a covalent bond between a nitrogen atom of the hydrophobic radial and a carbonyl of the copolyamino acid, thus forming an amide function originating from the reaction of an amine function of the precursor of the hydrophobic radical and an acid function borne by the precursor of the co-polyamino acid, or via a covalent bond between a carbonyl of the hydrophobic radical and a nitrogen atom in N-terminal position of the co-polyamino acid, thus forming an amide function originating from the reaction of an acid function of the precursor of the hydrophobic radical and an amine function in N-terminal position borne by the precursor of the co-polyamino acid; R is a radical selected from the group consisting of: a linear or branched divalent alkyl radical comprising 2 to 12 carbon atoms if GpR is a radical of formula II or 1 to 11 carbon atoms if GpR is a radical of formula II′ or II″; a linear or branched divalent alkyl radical comprising 2 to 11 carbon atoms if GpR is a radical of formula II or 1 to 11 carbon atoms if GpR is a radical of formula II′ or II″, 1 to 11 carbon atoms, said alkyl radical bearing one or more —CONH2 functions, and an unsubstituted ether or polyether radical comprising 4 to 14 carbon atoms and 1 to 5 oxygen atoms; A is a linear or branched alkyl radical comprising 1 to 6 carbon atoms; B is a linear or branched alkyl radical, optionally comprising an aromatic ring, comprising 1 to 9 carbon atoms; Cx is a linear or branched monovalent alkyl radical, in which x indicates the number of carbon atoms and: if p is equal to 1, x is from 11 to 25 (11≤x≤25): if p is equal to 2, x is from 9 to 15(9≤x≤15), the ratio i between the number of hydrophobic radicals and the number of glutamic or aspartic units being from 0 to 0.5 (0<i≤0.5); when several hydrophobic radicals are borne by a co-polyamino acid, then they are identical or different, the degree of polymerization DP of glutamic or aspartic units is from 5 to 250; the free acid functions being in the form of a salt of an alkali cation selected from the group consisting of Na+ and K+.
 8. The hydrophobic radical according to claim 7, wherein said hydrophobic radical is selected from the hydrophobic radicals of formula I in which p=1, represented by the following formula V:

in which GpR, GpA, GpC, r and a have the definitions given above.
 9. The hydrophobic radical according to claim 7, wherein said hydrophobic radical is selected from the hydrophobic radicals of formula I in which a=1 and p=2, represented by the following formula VI:

in which GpR, GpA, GpC and r have the definitions given above.
 10. A hydrophobic radical precursor Hy′ of the hydrophobic radical -Hy according to formula I′ as defined below:

in which GpR is a radical of formula II or II′:

GpA is a radical of formula III or III′:

GpC is a radical of formula IV:

the * indicate the sites of attachment of the different groups; a is a whole number equal to 0 or to 1; b is a whole number equal to 0 or to 1; p is a whole number equal to 1 or 2 and if p is equal to 1 then a is equal to 0 or to 1 and GpA is a radical of formula III′, and if p is equal to 2 then a is equal to 1, and GpA is a radical of formula III; c is a whole number equal to 0 or to 1, and if c is equal to 0 then d is equal to 1 or to 2; d is a whole number equal to 0, to 1 or to 2; r is a whole number equal to 0 or to 1, and if r is equal to 0 then the hydrophobic radical of formula I′ is attached to the co-polyamino acid via a covalent bond between a carbonyl of the hydrophobic radical and a nitrogen atom in N-terminal position of the co-polyamino acid, thus forming an amide function originating from the reaction of an amine function in N-terminal position of the precursor of the co-polyamino acid and an acid function borne by the precursor of the hydrophobic radical, and if r is equal to 1 then the hydrophobic radical of formula I′ is attached to the co-polyamino acid: via a covalent bond between a nitrogen atom of the hydrophobic radial and a carbonyl of the copolyamino acid, thus forming an amide function originating from the reaction of an amine function of the precursor of the hydrophobic radical and an acid function borne by the precursor of the co-polyamino acid, or via a covalent bond between a carbonyl of the hydrophobic radical and a nitrogen atom in N-terminal position of the co-polyamino acid, thus forming an amide function originating from the reaction of an acid function of the precursor of the hydrophobic radical and an amine function in N-terminal position borne by the precursor of the co-polyamino acid; R is a radical selected from the group consisting of: a linear or branched divalent alkyl radical comprising 2 to 12 carbon atoms if GpR is a radical of formula II or 1 to 11 carbon atoms if GpR is a radical of formula II′ or II″; a linear or branched divalent alkyl radical comprising 2 to 11 carbon atoms if GpR is a radical of formula II or 1 to 11 carbon atoms if GpR is a radical of formula II′ or II″, 1 to 11 carbon atoms, said alkyl radical bearing one or more —CONH2 functions, and an unsubstituted ether or polyether radical comprising 4 to 14 carbon atoms and 1 to 5 oxygen atoms; A is a linear or branched alkyl radical comprising 1 to 6 carbon atoms; B is a linear or branched alkyl radical, optionally comprising an aromatic ring, comprising 1 to 9 carbon atoms; Cx is a linear or branched monovalent alkyl radical, in which x indicates the number of carbon atoms and: if p is equal to 1, x is from 11 to 25 (11≤x≤25): if p is equal to 2, x is from 9 to 15 (9≤x≤15), the ratio i between the number of hydrophobic radicals and the number of glutamic or aspartic units being from 0 to 0.5 (0<i≤0.5); when several hydrophobic radicals are borne by a co-polyamino acid, then they are identical or different, the degree of polymerization DP of glutamic or aspartic units is from 5 to 250; the free acid functions being in the form of a salt of an alkali cation selected from the group consisting of Na+ and K+.
 11. The hydrophobic radical precursor Hy′ according to claim 10, wherein said hydrophobic radical precursor is selected from the hydrophobic radicals of formula I in which p=1, represented by the following formula V′:

GpR, GpA, GpC, r and a have the definitions given above.
 12. The hydrophobic radical precursor Hy′ according to claim 10, wherein said hydrophobic radical precursor is selected from the hydrophobic radicals of formula I in which a=1 and p=2, represented by the following formula VI′:

in which GpR, GpA, GpC and r have the definitions given above. 